Tunable interplay between exchange coupling and uniaxial magnetic anisotropy in epitaxial CoO/Au/Fe trilayers.

We show that the interaction between ferromagnetic Fe(110) and antiferromagnetic CoO(111) sublayers can be mediated and precisely tuned by a nonmagnetic Au spacer. Our results prove that the thickness of the Fe and Au layers can be chosen to modify the effective anisotropy of the Fe layer and the strength of the exchange bias interaction between Fe and CoO sublayers. Well-defined and tailorable magnetic anisotropy of the ferromagnet above Néel temperature of the antiferromagnet is a determining factor that governs exchange bias and interfacial CoO spins orientation at low temperatures. In particular, depending on the room temperature magnetic state of Fe, the low-temperature exchange bias in a zero-field cooled system can be turned "off" or "on". The other way around, we show that exchange bias can be the dominating magnetic anisotropy source for the ferromagnet and it is feasible to induce a 90-degree rotation of the easy axis as compared to the initial, exchange bias-free easy axis orientation.

Enterovirus 71 infection: report of an outbreak with two cases of paralysis and a review of the literature.

Enterovirus 71 (E-71) infection was first reported in 19745 in the United States; subsequent outbreaks were reported in worldwide distribution. In the summer of 1977, we identified 12 patients, mostly children, with E-71 infection. The striking feature of this outbreak is the occurrence of two cases with polio-like paralytic disease. Other diseases associated with E-71 included aseptic meningitis, meningoencephalitis, respiratory disease, gastroenteritis, and hand-foot-mouth disease. The spectrum of illness observed in our community was compared to that seen in other outbreaks. It is suggested that the significance of E-71 lies in its neuropathogenic potential.

Presentation of Yersinia enterocolitica enteritis in children.

Yersinia enterocolitica enteritis is a potentially treatable infection. To understand its seasonal incidence and clinical presentation in children, we reviewed case records of children seen in Cardinal Glennon Children's Hospital in St. Louis, MO. We found the incidence of Yersinia enteritis to be as frequent as enteritis caused by Campylobacter. It occurred more frequently during the winter months (P < 0.002) than during the rest of the year. Fever was common in infants with Yersinia enteritis. Abdominal pain and distention were infrequent. Seventeen (35%) patients were 3 months of age or younger; 4 of 17 (28%) developed Yersinia sepsis as a complication of the enteritis. Physicians should perform stool cultures for Y. enterocolitica in young infants who present with high fever and diarrhea in winter months, especially when there is blood in stools or the patient appears septic.

Effects of sonication and centrifugation of clinical specimens on the recovery of herpes simplex virus.

We have analyzed frozen and fresh specimens for herpes simplex virus (HSV) to determine the effect of sonication and centrifugation on virus recovery. After sonication, titers of 24/27 specimens increased from 1.3-30.8-fold with a mean increase of 6.8-fold. One hundred seventy-four fresh specimens were inoculated before and after sonication into CV-1 tube cultures. There was not a statistically significant difference in time to positivity between the sonicated and unsonicated portions. Thus, although sonication of specimens can sharply increase the viral titer of positive specimens, sonication of fresh specimens does not significantly enhance the isolation of HSV. To determine the effect of centrifugation of clinical specimens on recovery of HSV, thirty-one culture positive frozen specimens were centrifuged, and the supernatants and resuspended sonicated pellets were titered. Twenty-five specimens had sufficient recoverable virus to titer; in 21/25 (87.5%) specimens the titers were reduced in the supernatants after centrifugation (mean reduction 38%; range of 4-92%). Two of 31 (7%) supernatants were negative in culture while the sonicated pellets were positive. Thus, centrifugation of specimens prior to cell inoculation may compromise recovery of HSV.

Strategy for efficient detection of respiratory viruses in pediatric clinical specimens.

Direct immunofluorescence (IF) with a polyclonal respiratory syncytial virus (RSV)-specific antibody preparation was used for antigen detection during the 1982-1983 RSV season (155 specimens) and gave an overall sensitivity of 94% with 87% specificity compared with viral culture. Indirect IF was used in the 1983-1984 season (265 specimens) and exhibited sensitivity of 96% with specificity of 79%. During these two seasons, 42 of 224 (18.8%) specimens that were IF-negative for RSV grew viruses other than RSV. In the winter of 1984-1985, we screened 297 specimens for RSV by IF and 80 (27%) were positive. Forty-four (20%) of the IF-negative specimens were culture-positive for RSV(2) or other viruses(44). We conclude that, in the interest of cost reduction and expeditious detection of respiratory viruses, once a properly equipped laboratory has become thoroughly familiar with IF techniques, pediatric respiratory specimens can be screened for RSV by IF and only the IF-negative specimens need be inoculated into cell cultures for isolation of virus during the winter respiratory season.

Diagnosis of Trichomonas vaginalis in adolescent females: InPouch TV culture versus wet-mount microscopy.

PURPOSE: This study compared the InPouch TV culture to wet-mount, Diamond's culture medium, and Papanicolaou (Pap) smear for the diagnosis of trichomonas infection in sexually active adolescents. METHODS: A total of 467 subjects were recruited among 12-18-year-old girls who received pelvic examinations at two urban adolescent clinics. All girls were tested by wet-mount and InPouch TV. In addition 339 of 467 had cultures in Diamond's medium and 366 of 467 had Pap smears. Specimens were collected for InPouch TV and Diamond's cultures and read at 24-48 h and 5 days, and in the case of Diamond's cultures, also at 7 days. In a subset of subjects (268 of 467) who had all four tests done, sensitivities and specificities were calculated using Diamond's culture as the "gold standard." RESULTS: In the 467 subjects, 73 (15.6%) tested positive for trichomonas by at least one method. In the subset with all four tests done, sensitivities of the wet-mount and InPouch TV were 36% and 81%, respectively; while that of the Pap smear was 56%. The culture media were equally efficient in identifying Trichomonas vaginalis. There were no differences found between subjects with or without trichomonas infections in gynecological symptoms, previous history of sexually transmitted diseases, or use of a condom at last intercourse. CONCLUSIONS: InPouch TV culture is a good diagnostic method for T. vaginalis because of its long shelf-life, relatively low expense, and high sensitivity (over twice as sensitive as wet-mount).

Simplified method for typing herpes simplex virus by restriction endonuclease analysis.

We have developed a simplified method for unambiguously typing herpes simplex virus. The method depends on the production of cell-associated virus at 34 degrees C and subsequently, on the separation of cellular DNA and viral DNA by Dounce homogenization and the removal of nuclei by centrifugation. Viral nucleic acid was prepared from the cytoplasmic fraction and analyzed after restriction endonuclease cleavage. The method obviates the use of radioactive isotopes, and the viral DNA is effectively free of interfering cellular DNA.

Detection of rotavirus by hybridization with a nonradioactive synthetic DNA probe and comparison with commercial enzyme immunoassays and silver-stained polyacrylamide gels.

Three enzyme-linked immunosorbent assays (ELISAs) (rotavirus enzyme immunoassay [EIA; International Diagnostic Laboratories], Pathfinder [Kallestad Laboratories], and Rotaclone [Cambridge Bioscience, Inc.]) and hybridization of viral RNA with a nonradioactive, synthetic oligonucleotide DNA probe (SNAP; Molecular Biosystems, Inc.) were compared with silver-stained polyacrylamide gel electrophoresis (PAGE) of viral RNA for the detection of rotavirus in fecal specimens. Hybridization was performed following column purification of the viral RNA. Of 286 specimens analyzed by PAGE, SNAP, rotavirus EIA, Pathfinder, and Rotaclone, 88 were positive by PAGE. All 88 specimens were also positive by the other four assays. Nine specimens that were positive by one or more of the assays were positive by blocking ELISAs but were negative by PAGE. If these nine specimens were considered to be true positives, the final sensitivities and specificities were as follows: PAGE, 91 and 100%; SNAP, 94 and 97%; rotavirus EIA, 96 and 97%; Pathfinder, 100 and 94%; and Rotaclone, 96 and 97%, respectively.

Problems associated with the use of (E)-5-(2-bromovinyl)-2'-deoxyuridine for typing herpes simplex virus.

When 0.5 microgram of (E)-5-(2-bromovinyl)-2'-deoxyuridine (BVdU) per ml was incorporated directly into cell culture medium inoculated with eight known positive specimens, one herpes simplex virus type 1 (HSV-1) isolate grew in the presence of BVdU and was misidentified. By plaque assay, the titers of 15 HSV-1 strains were reduced by more than 3 log10 by BVdU, and the titers of 16 HSV-2 strains were reduced by less than 2 log10. Titers of HSV-1 acyclovir-resistant strains were reduced by less than 1.5 log10, which was characteristic of HSV-2 strains. Thus, typing of HSV isolates in the presence of BVdU by plaque assay is reliable only if information regarding previous antiviral therapy is obtained.

Sequence of an influenza virus hemagglutinin determined directly from a clinical sample.

The sequence of the HA1 region of the hemagglutinin gene of an influenza virus has been determined without growing the virus in eggs or in cultured cells. The virus used was an H1 strain of influenza A from a clinical specimen taken from a patient in 1987. RNA was extracted directly from virus that had been sedimented out of the transport medium in which the sample had been stored. DNA copies of the hemagglutinin gene, obtained by reverse transcription, were then amplified by the polymerase chain reaction and were sequenced by the dideoxy termination method. The deduced amino acid sequence is highly similar to that of other H1 viruses that had been isolated at about the same time and cultured for a limited number of passages in eggs. Furthermore, the HA1 sequence of progeny virus from this isolate obtained after one passage in chicken embryos is identical to that of the virus obtained directly from the nasopharynx. The results suggest that H1 isolates that have been grown for a limited number of passages in embryonated eggs have HA1 subunits that faithfully represent the virus population in the clinical samples from which they were derived.

Enhanced isolation of respiratory syncytial virus in cell culture.

During two winter seasons, we found that the combination of WI-38 or MRC-5 human lung fibroblasts plus primary rhesus monkey kidney (RhMK) and HEp-2 cell cultures yielded maximal isolation of respiratory syncytial virus. Cytopathic effects (CPE) developed earliest in RhMK cells and slowest in the human fibroblast lines. In RhMK cells, 50% of ultimately positive cultures showed CPE in 5 days, and 90% of positive cultures showed CPE within 7 days during both respiratory syncytial virus seasons.

Isolation and characterization of a naturally occurring parainfluenza 3 virus variant.

A parainfluenza 3 virus variant which failed to react with parainfluenza 3 virus-specific monoclonal anti-bodies from two commercial sources was isolated from a 14-month-old boy. Analysis of the coding region of the hemagglutinin-neuraminidase gene identified 36 nucleotide changes and 4 amino acid changes compared with a consensus sequence derived from strains isolated from 1957 through 1983. Two unique amino acid changes occurred at positions 174 and 283, which are close to identified epitopes in the hemagglutinin-neuraminidase protein. Ongoing viral surveillance to detect variants is important, particularly in regard to vaccine development.

Evaluation of two immunofluorescence assays with monoclonal antibodies for typing of herpes simplex virus.

An indirect immunofluorescence assay and a direct immunofluorescence assay were evaluated for typing clinical isolates of herpes simplex virus (HSV). The indirect immunofluorescence assay (Electro-Nucleonics, Inc.) correctly identified 16 HSV type 2 (HSV-2) isolates, but failed to identify 4 of 14 HSV-1 isolates because of background fluorescence and instability of reagents. Forty-nine HSV-1 isolates were correctly typed by direct immunofluorescence assay (Kallestad Laboratories, Inc.), but 1 of 39 HSV-2 isolates did not react with the HSV-2 type-specific antibody conjugate.

Effect of influenza B virus on nutrient transport in cultured epithelial cells.

The possibility that influenza virus could induce changes in membrane permeability to nutrients ordinarily concentrated within the cell was examined. Madin-Darby canine kidney cells were infected with egg-grown influenza B virus at 37 degrees C and pH 7.4 (a condition in which influenza virus enters cells by endocytosis). Control cells were mock-infected with allantoic fluid from chick embryos. Transport of phosphate, 2-deoxyglucose, and alpha-aminoisobutyric acid was measured at various intervals, 0 to 10 hours after infection. Uptake of alpha-aminoisobutyric acid and phosphate by infected cells was inhibited at 2 hours as compared with controls, whereas at 6 to 10 hours, the uptake of all nutrients was higher in infected cells. Infected cells preloaded with phosphate or 2-deoxyglucose did not demonstrate increased release of these nutrients. Thus, the virally induced inhibition of uptake early in infection is not a consequence of loss of membrane integrity. Transport studies were also performed in cells with prebound virus exposed to pH 5.0 for 60 seconds at 37 degrees C and then incubated at pH 7.4, at 37 degrees C. Under these conditions, influenza A viruses are known to enter the cell membrane by fusing directly with it and to initiate cell to cell fusion as well. We demonstrated that influenza B virus also caused cell fusion under these conditions. In contradistinction to studies described above at pH 7.4, fused, infected cells demonstrated both marked release and diminished uptake of nutrients as compared with controls. We conclude that influenza B virus does have an effect on host cell membrane permeability; the type of effect seen is markedly influenced by factors known to determine mode of virus entry into the cell.

Evaluation of the Abbott TESTPACK RSV enzyme immunoassay for detection of respiratory syncytial virus in nasopharyngeal swab specimens.

The Abbott TESTPACK RSV assay (Abbott Laboratories, North Chicago, Ill.), a rapid (20-min) enzyme immunoassay, was compared with culture and direct immunofluorescence (DFA) of nasopharyngeal cells for the detection of respiratory syncytial virus (RSV) in nasopharyngeal swab specimens. Nasopharyngeal swab specimens, collected from 234 infants, were placed in viral transport medium. Portions of specimen in transport medium were used for each test. Of 234 specimens, 70 (30%) were culture positive, 103 (44%) were DFA positive, 107 (46%) were culture or DFA positive, and 112 (48%) were TESTPACK RSV positive. Of 19 specimens positive by TESTPACK RSV but negative by culture or DFA, 15 were positive by the blocking assay. A total of 122 specimens were culture, DFA, or blocking assay positive; TESTPACK RSV detected 108 specimens (sensitivity, 89%). The specificity, positive predictive value, and negative predictive value of TESTPACK RSV as compared with those of culture, DFA, and the blocking assay were 96, 96, and 89%, respectively. By comparison, the sensitivity, specificity, positive predictive value, and negative predictive value of combined culture and DFA were 88, 100, 100, and 88%, respectively. TESTPACK RSV is a rapid and reliable enzyme immunoassay for the direct detection of RSV antigen in nasopharyngeal swab specimens.

Multidose, live attenuated, cold-recombinant, trivalent influenza vaccine in infants and young children.

Twenty-two healthy infants and children received either cold-recombinant, trivalent influenza vaccine or placebo in a three-dose vaccine trial. Most (82%) who received vaccine were seronegative to all three vaccine strains (10(6) TCID50/dose each): A/Kawasaki/9/86 (H1N1), A/Los Angeles/2/87 (H3N2), and B/Yamagata/16/88. Vaccine was administered intranasally at time 0 and 2 and 4 months later. The vaccine was well tolerated and immunogenic when administered in a multidose regimen. The first dose stimulated antibody to H1, H3, and B in 59%, 94%, and 35% of vaccinees, respectively, by hemagglutination inhibition (HAI) or ELISA. After two doses of vaccine, 93%, 93%, and 80% had antibody by HAI or ELISA to H1, H3, and B, respectively. Most vaccinees (67%) responded to all three viruses after two doses of vaccine. The third dose contributed little to the vaccine's immunogenicity. Multidose trivalent influenza vaccine is safe and induces an immune response in most vaccinees after two doses.

Improved DNA hybridization method for detection of acyclovir-resistant herpes simplex virus.

A simplified DNA hybridization method was developed to detect acyclovir-resistant isolates of herpes simplex virus. Herpes simplex virus-infected cell cultures in microtiter plates were treated with concentrations of acyclovir ranging from 8 to 0.015 micrograms/ml. At 48 h postinfection, infected cells were lysed by a one-step procedure and lysates were absorbed to membranes. Without further treatment, membranes were hybridized by using a herpes simplex virus-specific radioiodinated probe. The membranes were then washed and counted in a gamma counter. The elapsed time for assay performance was 4 h. Parallel plaque reduction assays were performed for comparison. The mean 50% inhibitory dose of in vivo- and in vitro-derived acyclovir-resistant, thymidine kinase-negative isolates was greater than 2 micrograms/ml by DNA hybridization. The 50% inhibitory dose of acyclovir-susceptible, thymidine kinase-positive isolates ranged from 0.01 to 1.1 micrograms/ml. This assay is simple and objective and should facilitate antiviral susceptibility testing in diagnostic laboratories.

Simultaneous detection and identification of human parainfluenza viruses 1, 2, and 3 from clinical samples by multiplex PCR.

Reverse transcription (RT)-PCR assays have been widely described for use in the diagnosis of human parainfluenza viruses (HPIVs) and other respiratory virus pathogens. However, these assays are mostly monospecific, requiring separate amplifications for each HPIV type. In the present work, we describe multiplex RT-PCR assays that detect and differentiate HPIV serotypes 1, 2, and 3 in a combined reaction. Specifically, a mixture of three pairs of primers to conserved regions of the hemagglutinin-neuraminidase gene of each HPIV serotype was used for primary amplification, yielding amplicons with similar sizes. For typing, a second amplification was performed with a mixture of nested primers, yielding amplicons with sizes easily differentiated by agarose gel electrophoresis. A modified single-amplification RT-PCR assay with fluorescence-labeled nested primers, followed by analysis of the labeled products on an automated sequencing gel, was also evaluated. Fifteen temporally and geographically diverse HPIV isolates from the Centers for Disease Control and Prevention archives and 26 of 30 (87%) previously positive nasopharyngeal specimens (8 of 10 positive for HPIV serotype 1 [HPIV1], 9 of 10 positive for HPIV2, and 9 of 10 positive for HPIV3) were positive and were correctly typed by both assays. Negative results were obtained with naso- or oropharyngeal specimens and/or culture isolates of 33 unrelated respiratory tract pathogens, including HPIV4, enterovirus, rhinovirus, respiratory syncytial virus, adenovirus, influenza virus, and Streptococcus pneumoniae. Our multiplex RT-PCR assays provide sensitive, specific, and simplified tools for the rapid diagnosis of HPIV infections.

Herpes simplex virus resistant to acyclovir. A study in a tertiary care center.

STUDY OBJECTIVE: To determine the sensitivity of herpes simplex virus isolates to acyclovir and the importance of resistant isolates in hospitalized patients. DESIGN: Retrospective incidence cohort study. SETTING: All herpes simplex virus isolates cultured over 1 year from patients followed at a tertiary care center. PATIENTS: Consecutive herpes simplex virus isolates were collected from 207 patients, including immunocompetent patients, patients with malignancy, neonates, bone marrow and organ transplant recipients, and patients seropositive for human immunodeficiency virus. MEASUREMENTS AND MAIN RESULTS: A rapid nucleic acid hybridization method was used to assess susceptibility to acyclovir. Acyclovir-resistant herpes simplex viruses were recovered from 7 of 148 immunocompromised patients (4.7%) but from none of 59 immunocompetent hosts. Clinical disease was found in all 7 patients with resistant herpes simplex virus and was more severe in pediatric patients. All resistant isolates were from acyclovir-treated patients and had absent or altered thymidine kinase activity by plaque autoradiography. CONCLUSION: Herpes simplex virus resistant to acyclovir arises relatively frequently in immunocompromised patients and may cause serious disease. Rapid detection of resistance permits antiviral therapy to be individualized. Antiviral susceptibility testing to monitor viral resistance should be encouraged, especially in tertiary care settings.

Impetigo contagiosa III. Comparative efficacy of oral erythromycin and topical mupirocin.

Ninety-seven patients with impetigo were prospectively enrolled in a study to determine the comparative efficacy of systemic and topical antibiotic therapy. After obtaining a bacterial culture from a representative lesion, the children were randomized to receive seven days of either oral erythromycin or topical mupirocin administered three times daily. Staphylococcus aureus alone was isolated from 51% and in association with group A beta-hemolytic streptococci (GABS) from 29%; GABS alone was isolated from 4% of patients. Of 48 children who received erythromycin, 43 (90%) were clinically improved or cured, and 11 of 17 were bacteriologically cured. Of 49 children who received mupirocin, 47 (96%) were clinically improved or cured, and 10 of 14 were bacteriologically cured. At three-week follow-up, clinical cure rates and number of secondary household cases of impetigo were equivalent in both treatment groups. Mupirocin appears to be a well-tolerated, albeit expensive, alternative to erythromycin for the treatment of impetigo.