Water-to-air transfer of virus.

Bubbles rising through suspensions of the bacteriophages T2 and T4 and of Escherichia coli adsorb and eject these particles in droplets that are formed when the bubbles burst. The concentration of the viruses in ejected droplets, determined from electron microscopy, exceeded the suspension concentration by 50 times. Similar results were obtained for Escherichia coli. The viability of some of the adsorbed particles was established by biological counts.

Virus transfer from surf to wind.

Bubbles in the sea surf adsorb and carry viruses to the surface where they are propelled into the air on tiny jets of seawater when the bubble bursts. The ejected jets become tiny drops of aerosol. The buble adsorption and virus concentration in the surf is analagous to industrial bubble levitation processes that concentrate metallic ores, enzymes, and finely divided organic crystals. Bubble levitation of viruses delibrately injected into the surf produced 200 times more virus per milliliter in the aerosol than were present in samples from the surf. Some aerosol drops created by the surf and carried by the wind fall out on the beach. The frequency of virus-bearing drops, that is, the number of plaques on seeded plates exposed on the beach, decreased exponentially with the distance downwind from the surf.

Molecular basis of age-dependent gastric inactivation of rhesus rotavirus in the mouse.

Rotavirus requires specific proteolytic activation by trypsin for efficient replication in tissue culture. To observe the nature of intestinal proteolytic activation of rotavirus in vivo, metabolically labeled rhesus rotavirus (RRV) grown in the presence of trypsin inhibitors was administered to adult and 10-d-old suckling mice by gavage. In the adult stomach, vp4 was cleaved in a manner distinct from in vitro trypsin cleavage. In the suckling stomach, RRV vp4 remains largely uncleaved. The alternative cleavage in the adult stomach was associated with a profound decrease in viral infectivity. vp4 from RRV recovered from the suckling small intestinal lumen was cleaved in a pattern similar or identical to in vitro trypsin-activated virus with bands comigrating with vp5* and vp8*. In contrast, vp4 was not observed in any recognizable form in RRV recovered from adult intestines. Comparison of infectivity of virus recovered from suckling and adult intestines revealed a 10,000-fold decrease in titer in the virus recovered from the adult intestine. In vitro digestions of RRV revealed that pepsin digestion can cleave RRV vp4 and markedly enhance acid-induced loss of rotavirus infectivity. Subsequent digestion with chymotrypsin removes most of the pepsin cleavage products of vp4. Virus injected directly into jejunal loops of adult mice and virus administered orally to adult mice pretreated with antiacid drugs retained infectivity. These studies indicate the development of gastric acid and pepsin secretion may be an important host defense factor in rotavirus gastroenteritis.

Liposome-mediated transfection of intact viral particles reveals that plasma membrane penetration determines permissivity of tissue culture cells to rotavirus.

Rotaviruses are an important cause of gastroenteritis in human infants. In vivo, rotavirus displays striking cell tropism with viral replication generally restricted to the villus tip enterocytes of the small intestine. We studied a panel of cell lines that vary significantly in their permissivity to rotavirus infection. L cells and HEp2 cells were relatively resistant to rotavirus infection compared with permissive Ma104 cells and HT29 cells. RNA transcription among the cell lines was proportional to antigen synthesis making a translational or posttranslational block an unlikely source of observed differences in susceptibility. All of the cell lines bound and internalized radiolabeled virus equally well, as measured by escape from surface protease treatment. Analysis of the escape of cell bound virus from neutralizing monoclonal antibody revealed that rotavirus did not immediately enter an eclipse phase in nonpermissive cells, but was internalized in an infectious form for several hours, possibly sequestered within endocytic vacuoles. L cells and HEp2 cells were as permissive as Ma104 and HT29 cells when rotavirus infection was mediated by transfection of single- or double-shelled rotavirus particles with cationic liposomes (Lipofectin). Rotavirus cell tropism in tissue culture cells is determined by the ability of infecting virions to traverse the plasma membrane of the cells into the cytoplasmic compartment.

Pseudomonas hyperimmune globulin passive immunotherapy for pulmonary exacerbations in cystic fibrosis.

We studied the effect of an intravenously administered gamma globulin [Ps-ivIG] enriched fivefold over conventional ivIG for Pseudomonas aeruginosa lipopolysaccharide [PA LPS] antibodies on ten patients with cystic fibrosis [CF] aged 19-32 years during hospitalization for pulmonary deterioration. All were colonized with greater than or equal to 1 PA phenotype resistant to all antibiotics at the time of admission and they received 500 mg/kg Ps-ivIG intravenously as a single dose in addition to conventional treatment, including antibiotics and chest physiotherapy. No adverse effects occurred. Circulating immune complexes and complement levels remained unchanged from baseline. Serum levels of anti-PA LPS IgG, as measured by ELISA for eight PA LPS immunotypes, increased to 244 +/- 65% (mean +/- SE) of baseline levels 1 hour post-infusion (P less than 0.01), remained significantly elevated during a mean hospital stay of 17 days, and returned to near baseline by follow-up 4 weeks after hospital discharge. Plasma half-life and clearance values were similar to those of other subjects receiving conventional ivIG. Sputum PA density declined from 3.0 to 1.2 x 10(8) cfu/mL 1 week post-infusion (P approximately equal to 0.05), and returned to baseline at follow-up. Serum anti-PA opsonic activity increased after infusion (P less than 0.01), but returned to baseline by 72 hours. Clinical scores improved from admission to discharge (P less than 0.005) without decline at follow-up. Forced vital capacity [FVC] and forced expiratory volume in one second [FEV1] increased from admission to discharge (P less than 0.01 and P less than 0.05, respectively) without decline at follow-up. Using autologous historical control data, standard hospital therapy without Ps-ivIG resulted in no improvement in FVC or FEV1, and a subsequent decline in these parameters (P less than 0.05 for each) during a similar follow-up period. This occurred despite the fact that half the patients did not have antibiotic-resistant PA on the control admission. We conclude that Ps-ivIG is a safe adjunctive therapy for pulmonary exacerbations in moderately ill cystic fibrosis patients colonized with resistant PA, and may be associated with both greater and more prolonged improvement in pulmonary function than standard therapy alone.

Morphological changes in human head hair subjected to various drug testing decontamination strategies.

Morphological changes in hair subjected to decontamination protocols were evaluated by scanning electron microscopy (SEM) as part of the National Laboratory Certification Program's (NLCP) efforts to develop proficiency testing materials in support of Federal Workplace Drug Testing programs. Hair from five different donors was evaluated. Hair samples were subjected to three decontamination protocols: (1) aqueous phosphate buffer, (2) methanol or (3) methylene chloride as models for aqueous, alcohol and polar organic solvent protocols, respectively. Under these protocols, samples of hair were treated for 225 min (aqueous), 15 min (alcohol), or 15 min (polar organic). After decontamination, hair strands were sputter coated with gold/palladium (AuPd) and observed by SEM. Modest lifting of cuticle scales was observed in hair treated with methanol and methylene chloride consistent with some changes to the cell membrane complex (CMC) between cuticle scales. Damage resulting from aqueous buffer treatment ranged from substantial degradation to apparent complete loss of cuticle scales. Fracture structures consistent with cuticle damage were also observed. Each decontamination protocol had a different impact on the cuticle of the hair shaft.

Computer-assisted research of medical records: a facilitated access file for physical examination, laboratory, and diagnostic data.

The Facilitated Access File, described herein, has proved to be an acceptable, relatively simple, yet feasible strategy for general medical record information retrieval at a university hospital clinic. Physicians record physical examination data in free-text mode, then through a rapid exercise of judgment, create an index to these data which is computer-stored. Such an index file provides future investigators facilitated access back to the original handwritten data and permits a variety of statistical studies. The system has been in use for over a year and has proved to be of value in clinical research and student teaching.

Murine intestinal mucins inhibit rotavirus infection.

BACKGROUND: Mucin, a population of polymeric glycoproteins, constitutes the primary component of the mucus layer that overlies the gastrointestinal tract. These studies aimed to determine whether murine intestinal mucins inhibit rotavirus infection. METHODS: Murine intestinal mucins were obtained by scraping segments of mouse intestine and purification via CsCl gradient centrifugation and sepharose 4B chromatography. Inhibition of infection was determined by quantitation of immunoperoxidase-stained cells after infection with mucin-rotavirus mixtures. RESULTS: Crude and purified intestinal mucins from suckling and adult mice are potent inhibitors of replication of a simian rotavirus, rhesus rotavirus (RRV), but weak inhibitors of other rotaviruses. In all preparations, colonic mucins were more potent inhibitors of RRV than small intestinal mucins. Suckling mucins neutralized RRV more effectively than adult mucins. In a panel of rotavirus reassortants, susceptibility to mucin inhibition correlated with the ability to hemagglutinate human type O erythrocytes and with RRV gene 4. Murine intestinal mucin inhibited RRV binding to MA104 cells, suggesting inhibition of virus-cell attachment to be the mechanism for neutralization. Mercaptoethanol or neuraminidase inhibited mucins' anti-RRV activities, implying the functional importance of mucins' polymeric structure and sialic acid content. CONCLUSIONS: These findings suggest that intestinal mucins represent a barrier to certain rotavirus infections.

Dansylcadaverine and cytochalasin D enhance rotavirus infection of murine L cells.

Although murine L cells bind and internalize rotavirus as well as permissive cell lines, L cells are essentially nonpermissive for rotaviruses. In nonpermissive cell lines such as L cells, internalized rotavirus fails to uncoat and remains as infectious, double-shelled particles. This block in the infectious cycle can be overcome by direct lipofection of viral particles into the L cell cytoplasm. We hypothesized that the internalized rotavirus particles within L cells are sequestered in the endocytic pathway and are unable to initiate infection. L cells were pretreated with a variety of inhibitors of endocytosis prior to infection with rhesus rotavirus. While agents which inhibit acidification of endosomes had no effect on rotavirus infection, two potential direct inhibitors of vesicular transport, dansylcadaverine and cytochalasin D, enhanced rotavirus infection of L cells 5- to 10-fold. All of the drugs, including both inhibitors of endocytosis and lysosomotrophic agents, significantly reduced infection of L cells by serotype 1 reovirus which is known to infect L cells by the endocytic pathway. Time course studies demonstrated that the drugs were effective in promoting rotavirus infection of L cells in only the early phases of infection. Pretreatment of L cells with dansylcadaverine significantly decreased the number of intact, double-shelled rotavirus particles sequestered within the cells. Inhibition of endocytosis may increase the efficiency of infection of L cells by rotavirus by allowing an increased proportion of attached rotavirus virions to enter cells by a productive route which is probably direct membrane penetration.

Acridine sensitivity of bacteriophage T2H in Escherichia coli.

Normally acridine-sensitive, Escherichia coli-T2H complexes are rendered acridine-resistant if the infecting bacteriophage mutant is either pr or q. If these pr or q mutants are treated to produce sensitive revertants, one obtains a mutation at any of several dye-sensitizing (ds) sites in the early enzyme region of the T2 map. The ds mutants are nonspecific suppressors because they reduce the resistance of complexes containing either pr or q to proflavine. The ds mutants are not identical in action, since some make pr or q sensitive to proflavine and quinacrine, and others, to proflavine alone. Two ds mutants have r to r(+) mutation patterns which differ, depending upon whether or not the ds is coupled with r7 (an rII mutant). The mutation patterns of r(+) to r are the same for both ds mutants and for wild type. We suggest that dye sensitization may consist of alterations of early enzymes so as to produce slightly different forms of deoxyribonucleic acid which are in turn dyesensitive.

Prevention of mast-cell degranulation by ketotifen in patients with physical urticarias.

The capacity for ketotifen to prevent mast-cell degranulation in vivo was studied in patients with physical urticarias. Patients were exposed to the appropriate stimulus to elicit their physical urticaria before and during ketotifen therapy. Histamine concentrations in plasma samples, obtained before and serially after the physical provocation, were determined by radioenzymatic thin-layer chromatography. Ketotifen therapy was associated with marked reductions in plasma histamine levels after stimulation and in clinical evidence of urticaria in each patient. A direct correlation of ketotifen therapy and a reduction in histamine release was confirmed in a patient with a cold-induced urticaria who was studied again after discontinuation and again after reinstitution of therapy. Although the mechanism of action is unknown, this report shows that ketotifen is capable of inhibiting cutaneous mast-cell degranulation and its accompanying symptoms. These findings suggest important therapeutic alternatives for patients with mast-cell-mediated diseases.

Iha: a novel Escherichia coli O157:H7 adherence-conferring molecule encoded on a recently acquired chromosomal island of conserved structure.

The mechanisms used by Shiga toxin (Stx)-producing Escherichia coli to adhere to epithelial cells are incompletely understood. Two cosmids from an E. coli O157:H7 DNA library contain an adherence-conferring chromosomal gene encoding a protein similar to iron-regulated gene A (IrgA) of Vibrio cholerae (M. B. Goldberg, S. A. Boyko, J. R. Butterton, J. A. Stoebner, S. M. Payne, and S. B. Calderwood, Mol. Microbiol. 6:2407-2418, 1992). We have termed the product of this gene the IrgA homologue adhesin (Iha), which is encoded by iha. Iha is 67 kDa in E. coli O157:H7 and 78 kDa in laboratory E. coli and is structurally unlike other known adhesins. DNA adjacent to iha contains tellurite resistance loci and is conserved in structure in distantly related pathogenic E. coli, but it is absent from nontoxigenic E. coli O55:H7, sorbitol-fermenting Stx-producing E. coli O157:H-, and laboratory E. coli. We have termed this region the tellurite resistance- and adherence-conferring island. We conclude that Iha is a novel bacterial adherence-conferring protein and is contained within an E. coli chromosomal island of conserved structure. Pathogenic E. coli O157:H7 has only recently acquired this island.

Characteristics of antibiotic-resistant Escherichia coli O157:H7 in Washington State, 1984-1991.

The resistance of Escherichia coli O157:H7 to amoxicillin/clavulanic acid, ampicillin, ceftazidime, ceftriaxone, cefuroxime, cephalothin, chloramphenicol, ciprofloxacin, gentamicin, streptomycin, sulfisoxazole, tetracycline, ticarcillin, tobramycin, and trimethoprim-sulfamethoxazole was examined, and resistant strains were characterized. All 56 isolates collected between 1984 and 1987 were susceptible to all antibiotics tested; 13 (7.4%) of 176 strains isolated between 1989 and 1991 were resistant to streptomycin, sulfisoxazole, and tetracycline. lambda-restriction fragment length polymorphism analysis suggested that the 13 resistant strains belonged to nine different clones. The emerging resistance of E. coli O157:H7 to antibiotics could portend an increased prevalence of this pathogen in food animals that receive antibiotics. Antimicrobial resistance of E. coli O157:H7 could be useful as a rapid epidemiologic marker and as a way to select this pathogen from suspected vehicles of transmission, but this resistance could also complicate therapeutic trials with sulfa-containing antibiotics.