Rapid presumptive identification of enteropathogenic Escherichia coli in faecal smears by means of fluorescent antibody: 1. Preparation and testing of reagents

In outbreaks of infantile diarrhoea caused by enteropathogenic Escherichia coli, rapid detection of the etiological agent is essential for specific and adequate therapy. Fluorescent antibody techniques have proved advantageous for rapid identification of many microorganisms. The development of fluorescein-labelled antibodies specific for nine serogroups of E. coli and the testing of these reagents for specificity are described in this paper. Results of testing pure cultures of various Enterobacteriaceae as well as faecal specimens from cases of diarrhoea are reported. These data show excellent correlation between the fluorescent antibody technique and conventional serological methods for identifying enteropathogenic types of E. coli. The immunofluorescent technique is more rapid than the usual bacteriological and serological methods in that smears from faecal specimens can be stained directly with the labelled antibodies. Details of the procedures for carrying out the technique are given in annexes.     

ORAL-MOTOR DYSFUNCTION AND FAILURE TO THRIVE AMONG INNER-CITY INFANTS

Non-organic failure to thrive among socio-economically deprived inner-city infants may be associated with abnormal oral-motor functioning and aspects of social adversity, such as disorganised meal-times. A Feeding Assessment Schedule was devised to rate oral-motor behaviour objectively and was used to test nine pairs of case and comparison children while they were being fed at home. Video-recordings were made for later analysis. The case infants had immature and abnormal oral-motor development that made them less able to be fed successfully. Temperamentally they were more 'difficult' than the comparison infants, and they were less adept at signalling their needs during meal-times. The case infants also were fed in inappropriate positions for their age, with more distractions and less suitable utensils.     

Erythrocyte Na+ and K+ transport systems in children with Bartter syndrome: increase in passive sodium permeability

Na+ and K+ intracellular content was studied in five children with Bartter syndrome and their age and race-paired controls. Na+ and K+ pump (ouabain sensitive) fluxes, Na+-K+ co-transport (furosemide sensitive), and rate constants of passive Na+ and K+ permeability were determined in each patient and control and also in six parents. The results show that in Bartter syndrome, there is a significant increase in the rate constant of passive Na+ permeability without any change in passive K+ permeability. This increase in the rate constant of passive permeability might explain at least partially the increased intracellular Na+ concentration also found in these patients. Moreover, the maximal rate of ouabain sensitive Na+ efflux was increased slightly, and co-transport fluxes were variable. Parents of patients had normal erythrocyte fluxes.     

Localization of deletion to a 300 Kb interval of chromosome 11q13 in cervical cancer

Previous molecular genetic studies on HeLa cell (a cervical cancer cell line) derived non-tumorigenic and tumorigenic hybrids have localized a tumor suppressor gene to the long arm of chromosome 11. Analysis of cervical cancer cell lines using chromosome 11 specific probes showed deletion and translocation of 11q13 sequences in five out of eight cell lines. Fluorescence in situ hybridization (FISH), using 11q13 specific probes, has shown interstitial deletion of 11q13 sequences in the HeLa cells. In order to determine whether 11q13 deletions occur in primary cervical tumors, we analysed 36 tumors using 20 different microsatellite and RFLP markers. Semi automated fluorescein based allelotyping was performed to identify loss of heterozygosity (LOH) in tumors. The results showed allelic loss in 17 (47%) tumors. Three different regions of loss, one near MEN1, the second near D11S913, and the third near INT2 locus were observed. The smallest region of deletion overlap at the D11S913 locus was localized to a 300 Kb distance between D11S4908 and D11S5023. Fluorescence in situ hybridization (FISH), using 11q13 specific cosmid and BAC (bacterial artificial chromosome) probes, confirmed allelic deletion in the tumors. PCR analysis further identified homozygous deletion of 11q13 sequences in a primary tumor, in HeLa cells and in two HeLa cell derived tumorigenic hybrid cell lines. The homozygous deletion in the cell lines was mapped to a 5.7 kb sequence of 11q13. We hypothesize therefore that a putative cervical cancer tumor suppressor gene exists within the 300 kb of chromosome 11q13.     

Insulin-like growth factor 1 gene (CA)n repeats and a variable number of tandem repeats of the insulin gene in Brazilian children born small for gestational age

OBJECTIVE:

To investigate the influence of (CA)n repeats in the insulin-like growth factor 1 gene and a variable number of tandem repeats of the insulin gene on birth size in children who are small or adequate-sized for gestational age and to correlate these polymorphisms with serum insulin-like growth factor 1 levels and insulin sensitivity in children who are small for gestational age, with and without catch-up growth.

PATIENTS AND METHODS:

We evaluated 439 infants: 297 that were adequate-sized for gestational age and 142 that were small for gestational age (66 with and 76 without catch-up). The number of (CA)n repeat in the insulin-like growth factor 1 gene and a variable number of tandem repeats in the insulin gene were analyzed using GENESCAN software and polymerase chain reaction followed by enzymatic digestion, respectively. Clinical and laboratory data were obtained from all patients.

RESULTS:

The height, body mass index, paternal height, target height and insulin-like growth factor 1 serum levels were higher in children who were small for gestational age with catch-up. There was no difference in the allelic and genotypic distributions of both polymorphisms between the adequate-sized and small infants or among small infants with and without catch-up. Similarly, the polymorphisms were not associated with clinical or laboratory variables.

CONCLUSION:

Polymorphisms of the (CA)n repeats of the insulin-like growth factor 1 gene and a variable number of tandem repeats of the insulin gene, separately or in combination, did not influence pre- or postnatal growth, insulin-like growth factor 1 serum levels or insulin resistance.