Virucidal activity of a new hand disinfectant with reduced ethanol content: comparison with other alcohol-based formulations.

A new formula with reduced ethanol content (55%) in combination with 10% propan-1-ol, 5.9% propan-1.2-diol, 5.7% butan-1.3-diol and 0.7% phosphoric acid exhibited a broad spectrum of virucidal activity. In quantitative suspension tests, with and without protein load, this formulation reduced the infectivity titre of seven enveloped (influenza A and B, herpes simplex 1 and 2, bovine corona, respiratory syncytial, vaccinia, hepatitis B, bovine viral diarrhoea) and four non-enveloped (hepatitis A, polio, rota, feline calici) viruses >10(3)-fold within 30s. In comparative testing, only 95% ethanol showed similar levels of activity. In fingerpad tests, the formulation produced a log10 reduction factor of the titre of poliovirus type 1 (Sabin) of 3.04 in 30s compared with 1.32 by 60% propan-2-ol. Testing against feline calicivirus produced a log10 reduction factor of 2.38 by the test formulation; in contrast, the log10 reduction factors with 70% ethanol and 70% propan-1-ol were 0.68 and 0.70, respectively.

Effects of propofol on the systolic and diastolic performance of the postischaemic, reperfused myocardium in rabbits.

BACKGROUND AND OBJECTIVE: The effect of propofol on myocardial dysfunction during ischaemia and reperfusion is controversial yet important because of its frequent use in cardiac anaesthesia. Although animal studies suggest a free radical-scavenging potential, the cardioprotective properties of propofol have not been demonstrated consistently in vivo. Previous studies focused on systolic function while diastolic function may be a more sensitive marker of ischaemic injury. The main aim was to document the effect of propofol on diastolic function in isolated, blood perfused rabbit hearts subjected to moderate global ischaemia and reperfusion. METHODS: Propofol 168 micromol L(-1), or the equivalent of its vehicle, Intralipid, was administered to 34 paced parabiotic Langendorff blood-perfused isolated rabbit hearts before and after 30 min of global normothermic ischaemia. Recovery of systolic function was quantified with the maximum rate of rise of left ventricular pressure. Diastolic performance was assessed using the time constant of the decline in left ventricular pressure (tau) and chamber stiffness (VdP/dV at 12 mmHg). RESULTS: Recovery of systolic function during reperfusion was comparable in the two groups. There was no difference in left ventricular pressure between the two groups at any time during the experiments. Chamber stiffness increased significantly during ischaemia and reperfusion in the control group (from 34 +/- 9 to 54 +/- 8 mmHg during ischaemia, and 43 +/- 5 mmHg after 30 min reperfusion; mean +/-95% confidence interval) but not in the propofol-treated group (29 +/- 5, 36 +/- 8 and 30 +/- 8 at baseline, ischaemia and 30 min reperfusion, respectively). CONCLUSIONS: Propofol has no protective effect on active relaxation or on systolic function in the present model, but it reduces ischaemic and postischaemic chamber stiffness.

Early diagnosis of herpes zoster by polymerase chain reaction.

BACKGROUND: Herpes zoster is a common disease caused by the varicella-zoster virus. The use of virostatic agents as early as possible is necessary in shortening zoster-associated pain. OBJECTIVES: Rapid diagnosis is necessary for the optimal efficacy of antiviral therapy. The diagnosis in the early stage of infection is often difficult. METHODS: In the present study skin biopsies of patients with herpes zoster and unclear skin changes were analysed by detecting viral DNA using the polymerase chain reaction (PCR) in order to amplify open reading frames (ORF) 14, 29 and 63. RESULTS: Varicella-zoster virus DNA could be detected with PCR of all three ORF not only from blisters but also from erythematous skin. CONCLUSIONS: PCR is the method of choice for the viral diagnosis in herpes zoster before blister eruption.

Hepatitis A virus: a test method for virucidal activity.

Hepatitis A virus (HAV) is closely related to the genus enterovirus. HAV is very stable and resistant to acid pH and elevated temperature, as well as to chemicals and environmental influences. Human poliovirus is still one of the model viruses for testing disinfectants but there are discussions about changing to hepatitis A virus. The purpose of this study was to develop a method for using adapted hepatitis A virus to test hand disinfectants. Using HAV strains HM175/24a and FRhK-4 cytopathic effects were visible rarely, and not before 14 days. To verify virus growth in cells a RT-PCR was developed. Two disinfectants tested did not show the required virucidal activity to satisfy current German guidelines.

Transmission of viruses via contact in ahousehold setting: experiments using bacteriophage straight phiX174 as a model virus.

Contamination of the environment with pathogens is the prerequisite for contact infections. The aim of this study was to elucidate how viruses can be transmitted from a primary contact person to further individuals. Bacteriophage straight phiX174 was chosen as a model virus. In its stability straight phiX174 is comparable with the most resistant human pathogenic viruses, e.g. polio- or parvoviruses. About 10(7)pfu were applied to exposed contact points such as door handles or the hands of volunteers. After touching of these handles and common social contacts like hand shaking, re-isolation rates were determined from the hands of our test persons. Contaminated door handles and skin surfaces were found to be efficient sources for potential infection. At least 14 persons could be contaminated by horizontal spread, one after the other by touching the same door handle. Successive transmission from one person to another could be followed up to the sixth contact person. These results were confirmed under everyday life conditions in a flat shared by four students. The transmission could not be prevented by the usual standards of hand hygiene, practised in this household. straight phiX174 could be reisolated after 24h from the hands of all persons tested even after normal use and cleaning of their hands. This might be improved by the use of liquid soap dispensers.

Transcriptional pattern of varicella-zoster virus glycoproteins E and I.

Glycoproteins I (gI) and E (gE) of the Varicella-zoster virus are encoded by the neighbouring open reading frames 67 and 68. From earlier work it is known that both genes are transcribed into several transcript species which differ in size. From gI, three transcripts of 1.65, 2.7 and 3.6 kb and from gE, two transcripts of 2.15 and 3.6 kb in size are known. We present further northern analysis and show that these various transcript species appear in different amounts at different times after infection. 12 h after infection, transcripts of 1.65 kb (gI) and 2.15 kb (gE) were clearly detectable, whereas the other transcripts appeared later on. RT-PCR experiments using a set of seven different primers provided clear evidence that gI and gE are transcribed both, mono- and bicistronically with predominance on the respective monocistronic transcript.

Effect of clevidipine, an ultra-short acting 1,4-dihydropyridine calcium channel blocking drug, on the potency of isoflurane in rats and dogs.

Clevidipine is a new lipophilic, ultra-short acting 1, 4-dihydropyridine calcium channel antagonist for intraoperative use. Because sarcolemnal calcium currents are involved in the mechanism of action of anaesthetics we questioned whether clevidipine alters the potency of volatile anaesthetics. We studied the effects of clevidipine on minimal alveolar concentration and the time to awaken from isoflurane anaesthesia. Sprague-Dawley rats were anaesthetized with isoflurane, and were allocated to one of four treatments prior to minimal alveolar concentration determination: (a) saline control; (b) clevidipine 20 nmol kg-1 min-1; (c) clevidipine 40 nmol kg-1 min-1; (d) clonidine 1 microg kg-1 - positive control. Ten mongrel dogs were anaesthetized with isoflurane and received a continuous infusion of clevidipine 6 nmol kg-1 min-1 or the solvent (Intralipid 20% - control). After 2 h of steady-state anaesthesia, minimal alveolar concentration awake and the time to awakening were recorded. Clevidipine reduced minimal alveolar concentration to a small but statistically significant extent (from 1.40 +/- 0.16 control to 1.23 +/- 0.13 vol % with 20 nmol kg-1 min-1 and to 1.27 +/- 0.12 vol % with 40 nmol kg-1 min-1; mean +/- SD; P < 0.05) in rats. Clonidine reduced minimal alveolar concentration to 0.90 +/- 0.17 vol % (mean +/- SD; P < 0.05). Minimal alveolar concentration awake and the duration of sleep were not affected by clevidipine in dogs. Clevidipine mildly reduces minimal alveolar concentration of isoflurane but does not prolong the duration of sleep and does not affect minimal alveolar concentration awake after a 2-h steady-state isoflurane anaesthesia.

Transcription factor Sp1 is involved in the regulation of varicella-zoster virus glycoprotein E.

Varicella-zoster virus glycoprotein E (ORF 68) belongs to the group of late genes. It is a major component of the virion envelope and can be found complexed with glycoprotein I on the infected host cell surface. Glycoprotein E (gE) expression is activated by IE4 and IE62. Also, cellular transcription factors, like Sp1, are able to influence the gE expression. Performing quantitative reverse transcription-polymerase chain reaction, we found no decrease in Sp1 mRNA levels at different times post-infection, indicating that Sp1 mRNA evade virion host shutoff effects. In addition, the Sp1 protein was detectable in highly infected cells. Electrophoretic mobility shift assays have shown a binding of Sp1 to both GC elements within the gE-5'untranslated region (5'UTR). Additional shift assays have notified a binding of TATA box binding protein to the TATA box of the gE promoter, which is characterized by an untypical TATACA motif. Promoter-reporter constructs have been made using mutated variants of the gE-5'UTR as promoters. In transfection studies, we found that the TATA deletion, as well as inactivations of both GC boxes, reduced the basal activity of the promoter. A complete loss of activity did not become measurable until eliminating both GC elements and the TATA box, indicating that these cis-elements substitute for each other in initiation of transcription of the gE-5'UTR.

[Measuring stroke volume of the ascending aorta with an extravascular Doppler ultrasound probe in comparison with aortic thermodilution]. / Messung des Herzzeitvolumens an der Aorta ascendens mit extravaskulärer Doppler-Ultraschallsonde im Vergleich zur aortalen Thermodilution.

Currently, no reliable minimally invasive method of measuring cardiac output continuously in neonates and children undergoing cardiac surgery is available. An extravascular Doppler probe was used to measure cardiac output in 15 New Zealand White rabbits (average weight 3.5 kg, range 2.5-4.5 kg). The results obtained were compared with cardiac outputs determined using the aortic thermodilution principle. The mean cardiac outputs measured with the extravascular Doppler probe was 0.37 +/- 0.01 l/min as compared with 0.39 +/- 0.01 l/min with aortic thermodilution. Regression analysis revealed a close correlation (r = 0.973) between the two techniques. The extravascular Doppler techniques is an option for continuous and reliable cardiac output measurement in small animals used in surgical experiments (open chest models) and in neonates or children during surgical repair of complicated congenital heart conditions.

Subclinical reactivation of herpes simplex virus type 1 in the oral cavity.

Reactivation in the oral cavity either symptomatically (recrudescence) or without symptoms (recurrence) may contribute to the transmission of herpes simplex virus type 1 (HSV-1), especially in critical areas of exposure such as dentistry. In order to measure the frequency of HSV-1 reactivation, nested polymerase chain reaction (PCR) was performed on oral swabs collected from 30 healthy people over a period of 58-161 days. In total 19 of 25 (76%) seropositive people were PCR-positive at least once, 6 of these 19 (32%) had recrudescence and 13 (68%) had only asymptomatic reactivation. Frequencies of additional recurrences were higher in people showing symptomatic reactivation than in those who had only recurrences. Recrudescence is a risk factor for elevated levels of asymptomatic HSV-shedding. In most cases HSV-1 was detected only by nested PCR investigated by early onset of therapy or time span before sampling.

Acyclovir treatment prevents varicella-zoster virus replication in PBMC during viremia.

The most effective antiviral therapy of varicella and zoster has become acyclovir. Using polymerase chain reaction specific for VZV ORF 14, ORF 29, ORF 63 as well as nucleic acid sequence-based amplification (ORF 63, ORF 68) we tested PBMC of patients with VZV-associated diseases for the presence of viral DNA and RNA, respectively. In PBMC of patients treated with acyclovir neither DNA nor RNA was detectable already one day after the onset of therapy. In three blood sample pairs from zoster patients we were able to detect viral nucleic acid before but not after acyclovir treatment. These results confirm clinical and epidemiological data. It can be concluded that treatment with acyclovir prevents VZV replication in peripheral blood mononuclear cells.

Influence of different cellular transcription factors on the regulation of varicella-zoster virus glycoproteins E (gE) and I (gI) UTR's activity.

Varicella-zoster virus (VZV) glycoproteins E (ORF 68) and I (ORF 67) are members of late genes. They belong to the major components of the virion envelope and can be found on the host cell surface as well. To get further insights in the regulation of gE and gI expression, which are known to be activated by IE4 and IE62, we analysed the intergenic regions of ORF 66/67 and ORF 67/68, containing the putative promoters of gI and gE. We have mapped the transcriptional start site of gE and have identified an extensive set of eucaryotic cis-elements: typical TATA- and CAAT-motifs and further regulatory sequences to facilitate interaction with eucaryotic transcription factors. Reporter constructs have been made using the intergenic regions of ORF 66/67 and ORF 67/68 as promoter elements. In cis-trans interaction studies, an influence on the regulation of transcription and reporter gene expression of overexpressed transfactors, LAP/LIP, Sp1, YY1 and NF-E2 has become measurable. In addition, protein-DNA binding assays using both gE- and gI-intergenic regions and cellular extracts from different VZV-permissive cells have suggested a binding of a 32 and 18 kD protein. In conclusion, these data indicate an involvement of common cellular transcription factors in the regulation of VZV late gene expression.

Effects of tramadol on minimum alveolar concentration (MAC) of isoflurane in rats.

It has been suggested previously that tramadol increases central nervous system activity and 'lightens' anaesthesia with volatile agents. We assessed the effects of tramadol on the minimum alveolar concentration (MAC) of isoflurane in 56 Wistar rats, instrumented chronically with an arterial and central venous catheter. The MAC of isoflurane was determined using the tail clamp method under three conditions: (1) after injection of saline (control); (2) after administration of tramadol 10 mg kg-1 i.v.; and (3) after administration of morphine 1 mg kg-1 i.v. The studies were repeated after treatment with the antagonists naloxone or yohimbine. Tramadol and morphine both reduced the MAC of isoflurane from mean 1.38 (SEM 0.05)% to 1.22 (0.06)% and 1.17 (0.06)%, respectively (P < 0.05). Concomitant administration of yohimbine did not abolish this reduction in MAC. In contrast, after pretreatment with naloxone, tramadol (1.47 (0.04)%) or morphine (1.38 (0.07)%) did not cause a reduction in the MAC of isoflurane compared with controls (1.39 (0.06)%). We conclude that tramadol and morphine reduced the MAC of isoflurane to a small but significant extent. For both drugs, this effect was related to their action at opioid receptors.

No acute varicella-zoster virus replication in peripheral blood mononuclear cells during postherpetic neuralgia.

Patients suffering from postherpetic neuralgia (PHN) were investigated whether varicella-zoster virus (VZV) DNA or RNA could be detected in their peripheral blood mononuclear cells (PBMCs). Altogether 16 samples were tested by standard polymerase chain reaction (PCR) for open reading frame (ORF) 14 and ORF 29, standard and nested PCR for ORF 63, and isothermal transcription-based nucleic acid amplification (NASBA) for ORF 63 and ORF 68. By these methods neither VZV DNA nor VZV RNA could be detected. The obtained results are in contrast to those of other authors (Vafai et al., 1988; Mahalingam et al., 1995) but support the hypothesis of Bennett (1994) and Kost and Straus (1996) proposing that PHN is not caused by acute VZV replication but a consequence of neuronal damage accompanying replication of VZV in ganglia during zoster episodes.

Subclinical reactivation of varicella-zoster virus in immunocompromised and immunocompetent individuals.

The occurrence of subclinical reactivation of varicella-zoster virus (VZV) in peripheral blood mononuclear cells (PBMC) from immunocompetent subjects >60 years old without any signs of VZV-caused illnesses, and from immunocompromised patients was investigated. Altogether, 223 samples were tested by nested ORF 63 PCR assay. In addition, all positive samples were tested by ORF 14, ORF 29 and ORF 63 PCR assays, as well as by ORF 63 and ORF 68 nucleic acid sequence-based amplification assays. In 5 samples, VZV-specific DNA, but no transcripts, could be detected. Three of them belonged to the group of >60-year-olds, 1 was HIV positive, the other was being treated with chemotherapy. The results confirm the observation of other authors that subclinical reactivation occurs in both immunocompromised and healthy individuals. The failure to detect DNA in samples taken from 2 individuals several weeks later excludes a long-lasting infection of VZV in PBMC.

Characterization of viremia at different stages of varicella-zoster virus infection.

Varicella-zoster virus (VZV) viremia at different stages of infection was characterized. Different approaches were used, polymerase chain reaction (PCR), isothermal transcription based nucleic acid amplification (NASBA), and immunofluorescence to describe and quantitate viral infection of peripheral blood mononuclear cells (PBMC). In patients with acute varicella 200 to 5,000 copies of the viral genome in every 150,000 PBMC were found with quantitative competitive PCR (QCPCR). With NASBA, viral transcriptional activity was detected in these cells. RNA transcribed from the immediate early gene IE 63 as well as from the late gene 68 were found, indicating a productive infection. Glycoprotein gE specific immunofluorescence visualized by confocal laser scanning microscopy revealed that only 1 in 10,000 to 100,000 PBMC was infected. T and B lymphocytes as well as monocytes expressed viral protein on their surface. Similar results were obtained with PBMC from immunocompetent zoster patients. In some cases a transient viremia was found shortly after the onset of rash, although the viral load seemed to be lower than in patients with varicella. Examination of blood samples from 16 persons with postherpetic neuralgia (PHN) signs of viral replication in PBMC were not detected. In conclusion, the data suggest that VZV viremia is a frequent event in patients with varicella and zoster, but not in those with postherpetic neuralgia. Moreover, the results indicated that subclinical reactivations occur both in immunocompromised and immunocompetent individuals.

Detection of varicella zoster virus DNA in single trigeminal ganglia by polymerase chain reaction.

Single human trigeminal ganglia were studied for DNA sequences of varicella zoster virus immediate early gene 63 by PCR. Out of 24 trigeminal ganglia five were tested positive for VZV DNA by standard PCR (21%), in six more VZV DNA was detectable using nested PCR (46%).