Structural determinants of processing and secretion of the Haemophilus influenzae hap protein.

Haemophilus influenzae elaborates a surface protein called Hap, which is associated with the capacity for intimate interaction with cultured epithelial cells. Expression of hap results in the production of three protein species: outer membrane proteins of approximately 155 kDa and 45 kDa and an extracellular protein of approximately 110 kDa. The 155 kDa protein corresponds to full-length mature Hap (without the signal sequence), and the 110 kDa extracellular protein represents the N-terminal portion of mature Hap (designated Haps). In the present study, we examined the mechanism of processing and secretion of Hap. Site-directed mutagenesis suggested that Hap is a serine protease that undergoes autoproteolytic cleavage to generate the 110 kDa extracellular protein and the 45 kDa outer membrane protein. Biochemical analysis confirmed this conclusion and established that cleavage occurs on the bacterial cell surface. Determination of N-terminal amino acid sequence and mutagenesis studies revealed that the 45 kDa protein corresponds to the C-terminal portion of Hap, starting at N1037. Analysis of the secondary structure of this protein (designated Hap beta) predicted formation of a beta-barrel with an N-terminal transmembrane alpha-helix followed by 14 transmembrane beta-strands. Additional analysis revealed that the final beta-strand contains an amino acid motif common to other beta-barrel outer membrane proteins. Upon deletion of this entire C-terminal consensus motif, Hap could no longer be detected in the outer membrane, and secretion of Haps was abolished. Deletion or complete alteration of the final three amino acid residues had a similar but less dramatic effect, suggesting that this terminal tripeptide is particularly important for outer membrane localization and/or stability of the protein. In contrast, isolated point mutations that disrupted the amphipathic nature of the consensus motif or eliminated the C-terminal tryptophan had no effect on outer membrane localization of Hap or secretion of Haps. These results provide insight into a growing family of Gram-negative bacterial exoproteins that are secreted by an IgA1 protease-like mechanism; in addition, they contribute to a better understanding of the structural determinants of targeting of beta-barrel proteins to the bacterial outer membrane.

Isolation and characterization of the Haemophilus influenzae uvrA gene.

The uvrA gene Haemophilus influenzae (Hi) was cloned and sequenced. Analysis of the deduced amino acid sequence revealed 81% identity and 90% similarity with the Escherichia coli UvrA protein. Consistent with a role of Hi uvrA in DNA repair, a Hi uvrA mutant exhibited increased sensitivity of UV irradiation. Furthermore, Hi uvrA was able to complement a mutation in the E. coli uvrA locus.

A Haemophilus influenzae IgA protease-like protein promotes intimate interaction with human epithelial cells.

Haemophilus influenzae represents a common cause of human disease and an important source of morbidity and mortality. Disease caused by this organism begins with colonization of the upper respiratory tract. Several studies indicate that H. influenzae is capable of binding to and entering cultured human cells, properties which are potentially of relevance to the process of colonization. In the present study, we isolated an H. influenzae gene designated hap, which is associated with the capacity for in vitro attachment and entry. Analysis of the derived amino acid sequence of hap demonstrated significant homology with the serine-type IgA1 proteases expressed by H. influenzae and Neisseria gonorrhoeae. It is notable that the hap product shares the catalytic domain of the IgA1 proteases and appears to be processed and secreted in an analogous manner. We speculate that the hap gene product is an important determinant of colonization, perhaps enabling the organism to evade the local immune response and thereby persist within the respiratory tract.

Diarrhea associated with Aeromonas species in children in day care centers.

Outbreaks of diarrhea caused by enteropathogens have been reported in day care centers (DCC), but Aeromonas species have not been implicated. This study evaluated 381 children involved in 51 outbreaks in four DCC to determine the association of Aeromonas species with diarrhea and to characterize the isolates. The organism was identified in two outbreaks of diarrhea. In one, Aeromonas species were isolated from 6 (24%) of 25 children and in the other from 5 (21%) of 24 children. Seven other Aeromonas strains from children in DCC were studied. Fourteen (78%) of 18 were Aeromonas caviae and 15 were from children with diarrhea. Of the isolates, 75% did not have plasmids detected; all others had unique plasmid patterns. All strains had different DNA content. Twenty-two control isolates of Aeromonas from children with diarrhea in Mexico and Dallas had different chromosomal DNA patterns. Most Aeromonas infections were associated with symptoms. Chromosomal DNA patterns differentiated Aeromonas strains better than did plasmid DNA patterns. The outbreaks of diarrhea were unusual in that several different Aeromonas genospecies were involved in each outbreak.

[Detection of gestational diabetes with the 50-gram glucose test: prevalence and relationship with to factors]. / Detección de la diabetes gestacional con la prueba de 50 gramos de glucosa: prevalencia y relación con factores de riesgo.

Prevalence of gestational diabetes was investigated in 222 pregnant women between the 24th and 28th weeks of gestation. A glucose screening test (GST) was performed with a 50 g glucose load, followed by a blood sample 1 hour later. Patients with glycemic levels greater than or equal to 140 mg/dl (7.8 mmol/l) 1 hour after the glucose administration were considered as having a positive GST and scheduled for a full oral glucose tolerance test (OGTT), which was interpreted according to O'Sullivan and Mahan criteria. Fifty-four patients had a positive GST, and 10 of them were diagnosed as having gestational diabetes mellitus. These figures indicate a gestational diabetes prevalence of 4.5% in general population and 18.5% in women selected by GST. The fetal weight index in patients with gestational diabetes was 1.13 +/- 0.10, whereas in normal women this index was significantly lower (p less than 0.01). Patients with positive GST and normal OGTT did not have an increased risk for fetal macrosomia. GST results were significantly higher in pregnant women with advanced age (greater than or equal to 35 years) and with late menarche (greater than or equal to 16 years), but did not vary according to maternal parity. Also, these results were significantly related with the number of risk factors for gestational diabetes encountered in a given patient, but not with any of them as examined alone, except for age. We conclude that GST performed in the 24th-28th weeks of gestation reveals 4.5% undetected gestational diabetes in our population. Certain maternal features (age, menarche, number of risk factors) influence on GST results.

[Psoriasis and chronic vitamin A poisoning]. / Psoriasis e intoxicación crónica por vitamina A.

A case of a five year old girl who during a treatment with high doses of A and D vitamins prescribed for a long time because "psoriasis" is showed. Clinical, radiologic and biochemical symptomatology of chronic poisoning by vitamin A is proved. Authors want to draw attention to dangers of long use of vitamin A.