Wide variation in effectiveness of laboratory disinfectants against bacteriophages.

AIMS: The purpose of this study was to identify an effective disinfectant for the inactivation of the bacteriophages (phages) being used in our laboratory, as published studies on phage inactivation are far from unanimous in their conclusions. METHODS AND RESULTS: The phages studied were three closely related strains of Myoviridae and three strains of Siphoviridae. Three disinfectants which are used commonly in microbiology laboratories were evaluated: Virkon (1%), ethanol (75%) and sodium hypochlorite (2500 ppm available chlorine). The most effective of these was Virkon, which inactivated all six phages rapidly. Ethanol was effective against the Myoviridae but had little effect on the Siphoviridae. Sodium hypochlorite was the least effective of the disinfectants evaluated. CONCLUSIONS: The findings of this study demonstrate a wide diversity in the effectiveness of disinfectants tested for inactivation of phages. SIGNIFICANCE AND IMPACT OF THE STUDY: Of the disinfectants tested Virkon is the most suitable choice for those unable to carry out disinfection validation studies, or where a broad spectrum disinfectant against phages is required. All of the phages in this study showed resilience to inactivation by sodium hypochlorite, and therefore this disinfectant is an unwise choice for use against phage without first assessing its effectiveness.

Survival rate of airborne Mycobacterium bovis.

Despite years of study the principle transmission route of bovine tuberculosis to cattle remains unresolved. The distribution of pathological lesions, which are concentrated in the respiratory system, and the very low dose of Mycobacterium bovis needed to initiate infection from a respiratory tract challenge suggest that the disease is spread by airborne transmission. Critical to the airborne transmission of a pathogenic microorganism is its ability to survive the stresses incurred whilst airborne. This study demonstrates that M. bovis is resistant to the stresses imposed immediately after becoming airborne, 94% surviving the first 10 min after aerosolisation. Once airborne the organism is robust, its viability decreasing with a half-life of approximately 1.5 hours. These findings support the hypothesis that airborne transmission is the principle route of infection for bovine tuberculosis.

Attaching-effacing lesions associated with Escherichia coli O157:H7 and other bacteria in experimentally infected conventional neonatal goats.

Four conventionally reared goats aged 6 days were inoculated orally with approximately 10(10) colony-forming units (cfu) of a non-verotoxigenic strain of Escherichia coli O157:H7. All remained clinically normal. Tissues were sampled under terminal anaesthesia at 24 (two animals), 48 and 72 h post-inoculation (hpi). E. coli O157:H7 was cultured from the ileum, caecum, colon and rectum of all animals, but the number of bacteria recovered at these sites varied between animals. Attaching-effacing (AE) lesions associated with O157 organisms, as confirmed by immunolabelling, were observed in the ileum of one of the two animals examined at 24 hpi, and in the ileum, caecum and proximal colon of an animal examined at 72 hpi. E. coli O157 organisms were detected at > or =10(5) cfu/g of tissue at these sites. In addition, AE lesions associated with unidentified bacteria were observed at various sites in the large bowel of the same animals. Lesions containing both E. coli O157 and unidentified bacteria (non-O157) were not observed. Non-O157 AE lesions were also observed in the large bowel of one of two uninoculated control animals. This indicated that three (one control and two inoculated) animals were colonized with an unidentified AE organism before the commencement of the experiment. The O157-associated AE lesions were observed only in animals colonized by non-O157 AE organisms and this raises questions about individual host susceptibility to AE lesions and whether non-O157 AE organisms influence colonization by E. coli O157.

Naturally acquired attaching and effacing Escherichia coli in sheep.

In a series of experiments involving the inoculation of sheep with Escherichia coli O157:H7, and subsequent detailed histopathological examination of the intestinal mucosa, attaching-effacing (AE) lesions formed by elements of the natural flora were observed in 18% of animals. These incidental AE lesions typically were small and sparse, and were not associated with clinical disease. It was possible to identify further some of the lesional bacteria, revealing that E. coli O115 had formed lesions in one of the seven affected animals, and similarly E. coli O26 had formed some of the lesions in another. As AE strains, source flocks, housing and feed sources were diverse, a common source of lesion-forming bacteria appears to be unlikely. It is postulated that subclinical AE lesions are a mechanism of persistence of AE bacteria in sheep.

Catastrophic presentation of mitochondrial disease due to a mutation in the tRNA(His) gene.

The authors describe a patient who presented with headache, seizures, and severe cerebral edema in whom they identified a novel mutation in the mitochondrial (mt-) tRNA(His) gene. This G12147A transition is heteroplasmic, predicted to disrupt a highly conserved base pair, and segregates with the cytochrome c oxidase deficiency in single muscle fibers.

Molecular and functional effects of the T14709C point mutation in the mitochondrial DNA of a patient with maternally inherited diabetes and deafness.

A heteroplasmic T to C transition at nucleotide position 14709 in the mitochondrial tRNA glutamic acid (tRNA(Glu)) gene has previously been associated with maternally inherited diabetes and deafness (MIDD). To investigate the pathogenic mechanism of the T14709C mutation, we have constructed transmitochondrial cell lines by transferring fibroblasts mitochondria from a patient with the mutation into human cells lacking mitochondrial DNA (mtDNA) (rho degrees cells). Clonal cybrid cell lines were obtained containing various levels of the heteroplasmic mutation, or exclusively mutated or wild-type mtDNA. Measurement of respiratory chain enzymatic activities failed to detect a difference between the homoplasmic mutant and homoplasmic wild-type cybrid cell lines. However, a subtle decrease in the steady-state levels of tRNA(Glu) transcripts in some mutant clones. Our studies suggest that the T14709C mutation is insufficient to lead impairment of mitochondrial function in homoplasmic osteosarcoma cybrid clones, and that we cannot exclude that the T14709C mutation affects mitochondrial function by a yet unidentified mechanism.

Progressive mitochondrial disease resulting from a novel missense mutation in the mitochondrial DNA ND3 gene.

We describe a 42-year-old man who presented with a progressive history of epilepsy, stroke-like episodes, bilateral optic atrophy, and cognitive decline. Investigation of his muscle biopsy revealed a specific defect in complex I activity. Subsequent analysis of the mitochondrial genome identified a novel heteroplasmic T10191C mutation in the ND3 gene. The mutation was present at lower levels in blood from the patient and unaffected maternal relatives and is the first pathogenic mitochondrial DNA mutation in the ND3 gene to be described.

Molecular characterisation and expression of a wound-inducible cDNA encoding a novel cinnamyl-alcohol dehydrogenase enzyme in lucerne (Medicago sativa L.)

A lucerne (alfalfa, Medicago sativa) stem cDNA library was screened with a cinnamyl-alcohol dehydrogenase (CAD) cDNA probe from tobacco (Nicotiana tabacum cv. Samsun). Two distinctly different cDNA clones (54% identical) were isolated and identified as putative CAD-encoding cDNAs by comparison of their nucleotide sequences with those of CAD-encoding DNA sequences from other plant species. One of the cDNAs, MsaCad2, was found to be 99.4% identical at the nucleotide level to the previously isolated lucerne cad cDNA which encodes a CAD isoform involved in lignin biosynthesis. The other cDNA, MsaCad1, has not been reported previously in lucerne, and encodes a protein related to the ELI3 class of elicitor-inducible defence-related plant proteins. The MsaCad1- and MsaCad2-encoded proteins were expressed in Escherichia coli and CAD1 was shown to be active with a range of cinnamyl, benzyl and aliphatic aldehyde substrates, while CAD2 was specific for the cinnamyl aldehydes only. Each of the respective genes is present as one or two copies. The MsaCad1 gene is expressed most actively in stem and floral tissue, whereas MsaCad2 is most actively expressed in stem, hypocotyl and root tissue. In stem tissue, expression of both genes occurs predominantly in internodes 4 and 5 (from the apex). MsaCad2, in contrast to MsaCad1, is not significantly expressed in the top three internodes of the stem. Both MsaCad1 and MsaCad2 are wound-inducible, and the wound-responsiveness of each gene is modulated by salicylic acid.

Contributory and exacerbating roles of gaseous ammonia and organic dust in the etiology of atrophic rhinitis.

Pigs reared commercially indoors are exposed to air heavily contaminated with particulate and gaseous pollutants. Epidemiological surveys have shown an association between the levels of these pollutants and the severity of lesions associated with the upper respiratory tract disease of swine atrophic rhinitis. This study investigated the role of aerial pollutants in the etiology of atrophic rhinitis induced by Pasteurella multocida. Forty, 1-week-old Large White piglets were weaned and divided into eight groups designated A to H. The groups were housed in Rochester exposure chambers and continuously exposed to the following pollutants: ovalbumin (groups A and B), ammonia (groups C and D), ovalbumin plus ammonia (groups E and F), and unpolluted air (groups G and H). The concentrations of pollutants used were 20 mg m-3 total mass and 5 mg m-3 respirable mass for ovalbumin dust and 50 ppm for ammonia. One week after exposure commenced, the pigs in groups A, C, E, and G were infected with P. multocida type D by intranasal inoculation. After 4 weeks of exposure to pollutants, the pigs were killed and the extent of turbinate atrophy was assessed with a morphometric index (MI). Control pigs kept in clean air and not inoculated with P. multocida (group H) had normal turbinate morphology with a mean MI of 41.12% (standard deviation [SD], +/- 1. 59%). In contrast, exposure to pollutants in the absence of P. multocida (groups B, D, and F) induced mild turbinate atrophy with mean MIs of 49.65% (SD, +/-1.96%), 51.04% (SD, +/-2.06%), and 49.88% (SD, +/-3.51%), respectively. A similar level of atrophy was also evoked by inoculation with P. multocida in the absence of pollutants (group G), giving a mean MI of 50.77% (SD, +/-2.07%). However, when P. multocida inoculation was combined with pollutant exposure (groups A, C, and E) moderate to severe turbinate atrophy occurred with mean MIs of 64.93% (SD, +/-4.64%), 59.18% (SD, +/-2.79%), and 73.30% (SD, +/-3.19%), respectively. The severity of atrophy was greatest in pigs exposed simultaneously to dust and ammonia. At the end of the exposure period, higher numbers of P. multocida bacteria were isolated from the tonsils than from the nasal membrane, per gram of tissue. The severity of turbinate atrophy in inoculated pigs was proportional to the number of P. multocida bacteria isolated from tonsils (r2 = 0.909, P < 0.05) and nasal membrane (r2 = 0.628, P < 0.05). These findings indicate that aerial pollutants contribute to the severity of lesions associated with atrophic rhinitis by facilitating colonization of the pig's upper respiratory tract by P. multocida and also by directly evoking mild atrophy.

Exploratory and confirmatory factor analyses of the Brief Symptom Inventory among substance abusers.

The Brief Symptom Inventory (BSI) is a multidimensional measure of psychological and somatic distress that is used to obtain detailed symptom profiles. The BSI has been questioned regarding its ability to differentiate among its proposed nine dimensions, and the factor structure underlying the BSI has not been confirmed with substance abusers. Exploratory factor analyses were completed on substance abusers (ntotal = 453; nwomen = 121; nmen = 332); nine factors were not identified. Five models were then submitted to confirmatory factor analyses using an independent sample of substance abusers (ntotal = 456; nwomen = 127; nmen = 329). A one-factor model (i.e., global psychological distress) best represented the data. Implications for using the BSI in research and counseling are discussed.

Effect of ovalbumin aerosol exposure on colonization of the porcine upper airway by Pasteurella multocida and effect of colonization on subsequent immune function.

Seventy-three piglets were weaned at 1 week of age, randomly assigned to 10 groups (A to J), accommodated in stainless steel exposure chambers, and exposed continuously to a controlled environment containing aerosolized ovalbumin. The concentrations of ovalbumin dust were as follows (milligrams per cubic meter): A and F, 16.6; B and G, 8.4; C and H, 4.2; D and I, 2.1; E and J, 0. At weekly intervals, the pigs were bled via venipuncture and anesthetized for nasal lavage and tonsilar biopsies performed for subsequent bacteriologic analysis. At 2 weeks of age, the pigs in groups A to E were challenged with toxigenic Pasteurella multocida (10(8) CFU pig(-1)), and at 6 weeks of age, the pigs were euthanatized. At postmortem, the extent of turbinate atrophy was assessed on the snout sections by using a morphometric index. Exposure to aerial ovalbumin resulted in a dose-dependent increase in serum antiovalbumin immunoglobulin G (IgG; P < 0.001) and serum antiovalbumin IgA (P < 0.001). Exposure also caused a significant increase in the numbers of P. multocida organisms isolated from the upper respiratory tract (P < 0.001) and a corresponding increase in turbinate atrophy, as judged by the morphometric index (P < 0.001). Concurrent challenge with P. multocida and ovalbumin resulted in a significant decrease in both the IgG and IgA responses to ovalbumin (P < 0.001). These results show that ovalbumin exposure increases pig susceptibility to P. multocida colonization and that toxigenic P. multocida modifies the serum IgG and IgA responses to ovalbumin in the pig. Both of these effects may enhance the virulence of this respiratory pathogen and so influence the pathogenesis of atrophic rhinitis in pigs.

Effects of ammonia inhalation and acetic acid pretreatment on colonization kinetics of toxigenic Pasteurella multocida within upper respiratory tracts of swine.

Pigs reared in intensive production systems are continuously exposed to ammonia released by the microbial degradation of their excrement. Exposure to this gas has been shown to increase the severity of the disease progressive atrophic rhinitis by facilitating colonization of the pig's upper respiratory tract by Pasteurella multocida. The etiological mechanism responsible for this synergy was investigated by studying the colonization kinetics of P. multocida enhanced by ammonia and comparing them with those evoked by an established disease model. Three-week-old Large White piglets were weaned and allocated to five experimental groups (groups A to E). Pigs in groups A and B were exposed continuously to ammonia at 20 ppm for the first 2 weeks of the study. Pigs in group C were pretreated with 0.5 ml of 1% acetic acid per nostril on days -2 and -1 of the study. On day 0 all the pigs in groups A, C, and D were inoculated with 1.4 x 10(8) toxigenic P. multocida organisms given by the intranasal route. The kinetics of P. multocida colonization were established by testing samples obtained at weekly intervals throughout the study. The study was terminated on day 37, and the extent of turbinate atrophy was determined by using a morphometric index. The results of the study showed that exposure to aerial ammonia for a limited period had a marked effect on the colonization of toxigenic P. multocida in the nasal cavities of pigs, which resulted in the almost total exclusion of commensal flora. In contrast, ammonia had only a limited effect on P. multocida colonization at the tonsil. The exacerbation of P. multocida colonization by ammonia was restricted to the period of ammonia exposure, and the number of P. multocida organisms colonizing the upper respiratory tract declined rapidly upon the cessation of exposure to ammonia. During the exposure period, the ammonia levels in mucus recovered from the nasal cavity and tonsil were found to be 7- and 3.5-fold higher, respectively, than the levels in samples taken from unexposed controls. Acetic acid pretreatment also induced marked colonization of the nasal cavity which, in contrast to that induced by ammonia, persisted throughout the time course of the study. Furthermore, acetic acid pretreatment induced marked but transient colonization of the tonsil. These findings suggest that the synergistic effect of ammonia acts through an etiological mechanism different from that evoked by acetic acid pretreatment. A strong correlation was found between the numbers of P. multocida organisms isolated from the nasal cavity and the severity of clinical lesions, as determined by using a morphometric index. The data presented in the paper highlight the potential importance of ammonia as an exacerbating factor in respiratory disease of intensively reared livestock.

The ear. Excision and repair.

An ordinary looking ear is important for a person's overall appearance. A review of the auricular anatomy is advised prior to operating on the ear. Surgery on the ear is done for both benign and malignant lesions. Many surgical options exist for the repair of ear defects including simple excisions, wedge excisions, complex linear closures, flaps, and skin grafts. Although the procedure may be challenging, the results obtained are often excellent.

Expression patterns of three genes in the stem of lucerne (Medicago sativa).

We have identified three stem abundantly expressed genes in lucerne (alfalfa, Medicago sativa). A cDNA library, constructed from lucerne stem polyadenylated RNA, was screened by differential hybridization. From this screening, cDNA clones that correspond to genes which are preferentially, or specifically, expressed in the stem were isolated. MsaS1 encodes an unidentified protein, MsaS2 encodes an S-adenosyl-homocysteine hydrolase and MsaS3 encodes an extensin-like protein. Northern blot analysis of RNA isolated from individual stem internodes indicated that the three corresponding genes show differing developmental patterns of expression. The expression of MsaS1 was confined to the youngest stem tissue and may be regulated by sucrose. In stem tissue the level of RNA for the three genes decreased in response to wounding. Tissue print hybridization analysis was used to localize the expression of the genes to the xylem side of vascular bundles in lucerne stems.

Primary cutaneous meningioma associated with von Recklinghausen's disease.

Cutaneous meningeal tumors are rare and can pose a diagnostic problem. We present a case of a 12-year-old girl with a family history of von Recklinghausen's disease. The patient was asymptomatic until the age of 11, when she developed two lesions on the head, both diagnosed as plexiform neurofibroma. Subsequently, she presented with a subcutaneous nodule on the left posterior occipital scalp which was excised. On histology, the tumor was composed of spindle-shaped cells with diffuse and nested patterns. A whorled configuration of the cells, with occasional giant cells and psammoma bodies, was present. There was no evidence of connection between the tumor and the underlying tissues. Immunohistochemical studies were positive for epithelial membrane antigen, vimentin, and weakly for neuron-specific enolase. Cytokeratin, S-100 protein, and muscle markers were negative. Based on these features, the diagnosis of cutaneous meningioma was made. An MRI examination failed to detect any communication between the tumor site and the meninges; however, asymptomatic bilateral acoustic neuromas were identified. This case, besides being of interest as a primary cutaneous meningioma, also documents a unique combination of findings, i.e., plexiform neurofibroma, meningioma, and cerebellopontine acoustic neuromas, which should alert the clinician to a forme fruste presentation of von Recklinghausen's disease.

Recovery of heat-injured Listeria monocytogenes.

Concern has been expressed that the ability of heat-injured Listeria monocytogenes cells to resuscitate during refrigerated storage of food may lead to underestimation of their heat resistance. The recovery of heat-injured L. monocytogenes was therefore examined as a function of incubation temperature and composition of recovery medium. Heat-injured cells exhibited a broad optimum temperature for recovery centered around 20-25 degrees C. The best recovery medium of those tested was blood agar. Incubation of cells in broth or chicken slurry at 5 degrees C (cold-enrichment) did not allow repair of potentially lethal injury i.e. it did not allow recovery of cells that would otherwise have died if incubated at a higher temperature. In some cases incubation of heat-injured cells at 5 degrees C resulted in death of a proportion of the population. Repair of sublethal heat-injury, measured as the time of incubation in tryptone soya broth needed to regain the ability to grow on Listeria selective agar, was slower and less complete at 25 degrees C than at 2 degrees C; repair took 10-15 h at 25 degrees C compared with 8-12 days at 5 degrees C. Refrigeration of heat-treated foods should not therefore increase the risk that heat-injured cells will recover from the heat treatment.

Complications of cutaneous surgery.

There are many complications that may develop when surgical procedures are performed. With adequate preoperative preparation and careful technique, however, they can be kept to a minimum. Foreknowledge of the possible difficulties will enable the physician to recognize them when they are encountered and to provide appropriate treatment.

Fever in the elderly. Production of leukocytic pyrogen by monocytes from elderly persons.

In an attempt to explain the diminished febrile response of the elderly, we studied the first step in fever generation, that of production of leukocytic pyrogen (LP) by monocytes. Monocytes from 25 healthy elderly volunteers (ages 65-91) and 24 healthy young volunteers (ages 17-38) were stimulated with Staphylococcus epidermidis to release LP; LP activity in the culture supernatants was assayed by measuring the pyrogenic response in rabbits and rats and the fall in plasma iron and zinc in rats. Monocytes from elderly volunteers produced slightly less LP than monocytes from young volunteers, but the difference was not statistically significant. The amount of LP produced was not correlated with age. Therefore, the diminished febrile response of the elderly is not the result of an intrinsic defect in the monocyte's ability to make LP. Other explanations relating to the central effect of LP and the effector response to LP in the elderly should be sought.