[Evaluation of the Yersinia enterocolitica pathogenicity through some phenotypic and genotypic characters: CRMOX agar positivity and presence of the ail gene]. / Valutazione della patogenicità di Yersinia enterocolitica attraverso alcuni caratteri fenotipici e genotipici: positività su CRMOX agar e presenza del gene ail.

Sixty-nine strains of Y. enterocolitica isolated from environmental and human matrices (waste water, food and faeces) were studied in order to evidence the presence of ail gene, calcium-dependency and Congo Red absorption for pathogenic strains identification. Out of 24 clinical strains, the ail gene was present in 21 (87%), among which 79% were CRMOX-positive as well. On the contrary, none of the 45 environmental strains showed the ail gene although only one (isolated from cooked vegetables) was CRMOX agar positive. Our results confirmed the importance of molecular methods to evidence the Y. enterocolitica pathogenic strains. However, our study pointed also the utility to consider the approach of classic bacteriology, like the subcoltivation on CRMOX agar to show calcium-dependency and Congo Red absorption. In particular, when dealing with environmental isolates, that medium will be useful as a preliminary screening to identify those isolates which need further research to indicate their pathogenic potential by the use of more complex but also more expensive molecular methods.

Biometrical implications of factorial experiments for the study of lymphocyte mitogenic response.

Lymphocyte responsiveness to mitogens is affected by a number of experimental variables; the latter may be conveniently studied several at a time, following a factorial design that allows the complete evaluation of the effects of each variable and of their interplays. Statistical inferences may be drawn through an analysis of variance, provided that the data conform to the underlying model, i.e. that the effects are additive and that the errors are normally distributed and homoscedastic. Thymidine incorporation data should first be transformed to log cpm; the effects on the metameter may then be assumed to be additive, and their errors approach normality; the error variances may be affected by the experimental variables involved, so that the homogeneity of the error must be checked before relying on the pooled estimate of the error for parametric tests of significance. The above considerations have been illustrated with factorial experiments on [3H]TdR incorporation by rabbit spleen cells stimulated with varying amounts of PHA-P or PHA-M, under a variety of culture conditions.

Age-related changes in the prevalence of precipitating antibodies to BK virus in infants and children.

Nearly 1000 sera from children were tested by immunoelectro-osmophoresis against BK virus, and age-specific prevalence rates were estimated from birth until the age of 12 years. Declining rates during the first 12 months showed the waning of passive immunity, which at birth reflects the mother's immune status. The changes of prevalence suggested that the peak incidence of primary infections occurred at about 2 years, with an estimated peak annual rate of 24.6%.

Immunoelectroosmophoresis for the human papovavirus BK.

The reaction between BK virus and its antibodies may well be visualized with immunoelectroosmophoresis (IEOP). Serum antibody levels may be compared or graded by reacting them against a continuous antigen front. In seroepidemiologic surveys IEOP compares favourably with hemagglutination inhibition in terms of sensitivity, practicability and reproducibility. Technical conditions are described which are suitable for antigen detection, quantification and identification.

Outbreak of Clostridium difficile diarrhoea in an orthopaedic unit: evidence by phage-typing for cross-infection.

In a three-week period five patients had diarrhoea in an orthopaedic unit. The first case was clinically diagnosed as pseudomembranous colitis but the causative agent was not sought. Of the remaining cases, two were Clostridium difficile positive. The outbreak then apparently ceased, but during the following several days two of seven stool samples taken at random from asymptomatic patients yielded C. difficile. Phage-typing of the isolates showed that all apparently belonged to the same strain.

HPLC does not differentiate Mycobacterium paratuberculosis from Mycobacterium avium.

HPLC, which is gaining its place as identification tool in mycobacteriology laboratories, has been proposed to distinguish Mycobacterium paratuberculosis from Mycobacterium avium. We had reported no significant difference between M. avium and M. paratuberculosis reference strain ATCC 19698. Because of the advantages offered by such a method, we enlarged our observations to include more isolates of M. paratuberculosis. Within the double cluster of peaks obtained by both M. avium and M. paratuberculosis, we could not find a consistent difference typical of M. paratuberculosis. Therefore, the present study confirmed that M. avium and M. paratuberculosis could not be distinguished by HPLC, raising doubts of a straightforward use of HPLC to identify M. paratuberculosis.

Metal uptake in a coastal diatom influenced by major nutrients (N, P, and Si).

The influence of major nutrient additions on trace metal uptake (Cd, Se, and Zn) in a coastal diatom Thalassiosira pseudonana was investigated. A short-term exposure (5 h) was employed to measure the concentration factor (ratio of metal concentration in the cells to metal concentration in the medium) which was used as a kinetic parameter to quantify the relative rate of metal uptake in the cells. Following an initial rapid surface sorption, a linear pattern of uptake over time was typical for Cd, Se and Zn, indicating that these metals may have been transported intracellularly during the short-term exposure period. N addition significantly increased the rates of Cd uptake in the cells. Although the concentration factor of Zn increased with N addition, statistical analysis indicated that N addition did not significantly affect the rate of Zn uptake in the cells. Se uptake in the cells was independent of N additions, but was lowered with increasing Si concentration. Si addition did not significantly affect Cd and Zn uptake. Similarly, P addition did not influence the rates of metal uptake in the cells. There was a significant correlation between the rate of uptake of Cd and Zn and the cell growth rate. This study demonstrated that water quality brought about by a change in nutrient condition could considerably influence the uptake of metals by marine phytoplankton.

Influences of phosphate and silicate on Cr(VI) and Se(IV) accumulation in marine phytoplankton.

The influences of major nutrient additions (P, Si, or N) on the accumulation of chromate [Cr(VI)] and selenite [Se(IV)] in three species of marine phytoplankton, including two diatoms (Thalassiosira pseudonana, and Skeletonema costatum), and one green alga (Chlorella autotrophica) were examined. Accumulation rate was quantified by measurements of metal concentration factor in algal cells over a short 5-h exposure period. Our results suggested that Se(IV) was little bioavailable to C. autotrophica and S. costatum, but was highly bioavailable to diatom T. pseudonana. Se(IV) accumulation in both diatoms and green alga was inversely dependent on the ambient P and Si concentration. Concentration factor decreased by 2.4-8.1 and 1.5-4.6 times with the additions of 7.2 microM P or 105.6 microM Si, respectively, after 5 h of exposure. Bioconcentration factors of Se(IV) in these species determined under laboratory conditions, which generally employ high nutrient concentrations, may not be applicable to those prevailing in the field. In contrast to Se(IV), Cr(VI) accumulation in the green alga and two diatoms was not related to the ambient P concentrations, but was inversely dependent on the ambient Si concentrations. These data demonstrated that the influences of major nutrients on anionic metal accumulation were highly species-specific, both for metals and phytoplankton. Nutrient enrichments in many water bodies may potentially affect the accumulation of anionic metals, and thus their biological transport and fate in these environments.

Nontuberculous mycobacteria in hospital water systems: application of HPLC for identification of environmental mycobacteria.

Nontuberculous mycobacteria (NTM), ubiquitous in water environments, are increasingly recognized as nosocomial pathogens. Our study reports a one-year survey of the water system of two hospitals, A and B, in a small town near Florence, Italy. NTM were found throughout the study period in both settings, but B showed a significantly higher mycobacterial load. Mycobacterium gordonae and Mycobacterium fortuitum were the most frequent species isolated. Identification was carried out by conventional techniques and by high performance liquid chromatography (HPLC) analysis of cell wall mycolic acids. HPLC profiling could be used as a first-choice method for identification of environmental mycobacteria.

Betalactam therapy and intestinal flora.

Betalactams, mainly when orally administered, may lead to intestinal flora modifications related to their spectrum of activity, rate of absorption and degradation. therefore it is important to investigate the possible influence of recently developed oral cephem derivatives on normal human microflora. We have investigated the impact on normal human intestinal flora in a 10-day course with cefetamet-pivoxil (CET, 500 mg BID) in comparison to cefixime (CFX, 400 mg qD) or cefuroxime axetil (CA, 250 mg BID) in 24 patients suffering from acute exacerbation of chronic bronchitis. Stool specimens were taken before (day 0), at the end (day 10) and 14 days after treatment (day 24) and quali-quantitative microflora composition was determined with a detection limit of 10 CFU/g dry weight. Treatment with CET caused slight and non-significant modifications of normal intestinal flora. On the contrary CFX and CA significantly affect Enterobacteriaceae and clostridia with a concomitant increase in enterococci for CFX. With both CFX and CA there was a new appearance of Salmonella spp. as well as Clostridium difficile in 4 and 2 cases, respectively. Therefore CET seems to affect normal bowel flora minimally in comparison to other oral cephalosporins. This aspect might contribute to the low incidence of GI related side effects in patients treated with CEt for longer than 1 week.

In-vitro activity of nisin against clinical isolates of Clostridium difficile.

Nisin is a cationic peptide produced by Lactococcus lactis. Its activity against clinical isolates of Clostridium difficile was compared to that of vancomycin and metronidazole by minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and time-kill studies. Nisin was more active than the other agents, with a MIC90 of 0.256 mg/L and strong bactericidal activity. Nisin may be a promising agent for the management of C. difficile associated diarrhea.

Effects of major nutrient additions on metal uptake in phytoplankton.

We examined the influences of major nutrients (N, P, Si) on the accumulation of three trace metals [Cd, Se(IV), and Zn] in four species of marine phytoplankton (diatom, green alga, dinoflagellate, prasinophyte). Relative metal uptake was quantified by the kinetic measurements of metal concentration factor over a short exposure period. Our study demonstrated that nutrient addition significantly influenced the metal uptake rate and the cell growth rate in all four phytoplankton species. An increase in ambient N concentration considerably enhanced metal uptake by the cells. The dry weight concentration factor increased by 2.4-14.9 times for Cd, 1.1-4.0 times for Se, and 1.1-5.4 times for Zn in all four phytoplankton species with an addition of 176.4 microM N. The effects of P or Si addition on metal uptake and cell growth were less pronounced than the effects of N addition. Under most circumstances the rate of metal uptake increased exponentially with increasing cell growth rate constant. Only Se(IV) uptake in the diatom Phaeodactylum tricornutum was not correlated with cell growth rate. Se(IV) was not accumulated by the green algae Chlorella autotrophica at a high P concentration (7.2 microM), but appreciable accumulation was documented in cells inoculated without P addition. Our study therefore demonstrated that nutrient enrichments in many coastal waters can considerably affect trace metal uptake in phytoplankton and presumably metal trophic transfer in marine food chains.

[Proteins bound to satellite DNA, are present in human nuclear matrix cell preparations]. / Belok, sviazannyi s satellitnoi DNK, prisutstvuet v preparatakh iadernogo matriksa kletok cheloveka.

The presence of protein or complex of proteins that specifically bind to human satellite 3 (HS3) was shown during investigations of the nuclear matrix. The specificity of binding of HS3 was shown by using nuclear matrix immobilized on nitrocellulose. The activity disappeared after extractions of the nuclear matrix. The presence of specific activity in low salt extract was shown by gel retardation assay with whole HS3 fragment. All the subfragments of HS3 after Sau3A restriction (1 kb, 0.36 kb, 0.41 kb) also retarded in the mixture with this protein extract. DNA-protein complexes were stable even in the presence of a 1000-fold excess of competitive DNA. These data are discussed in the frame of hypotheses about the three dimensional organization of interphase chromatin.

[Ipsilateral central-type facial palsy and contralateral hemiparesis associated with unilateral medial medullary infarction: a case report].

Clinical pictures of medial medullary syndrome are variable, depending upon the extent of the lesion. Facial palsy has rarely been observed even in medullary infarction. However, central-type facial palsy is usually found contralaterally to the infarct area at the level of the rostral medulla. In the present report, we discuss the pathogenesis of the neurological manifestations in a 57-year-old man with hypertension. The patient presented with mild left facial palsy of central type, right hemiparesis, paresthesia, with deep sensory disturbance of the right extremities. An MRI of the brain showed an infarction localized in the medial region of the left upper medulla. Although the exact course of the supranuclear facial pathways remains controversial, the ipsilateral central facial palsy in this patient is considered to have two possible causes: the interruption of aberrant fibers of the corticobulbar tract, which branch off and swing back at the level of the upper-middle medulla, or the disruption of recurrent ascending fibers from the contralateral pyramidal tract, through decussation.

[Pseudomonas aeruginosa antibodies in patients with cystic fibrosis: clinical implications]. / Anticorpi anti-Pseudomonas aeruginosa in pazienti con fibrosi cistica: implicazioni cliniche.

Patients chronically colonised by P. aeruginosa develop a pronounced antibody response against P. aeruginosa that can be used to discriminate between superficial colonization and chronic infection. Anti-P. aeruginosa antibodies fail to afford protection against this pathogen; moreover high levels of antibodies are correlated with a poor prognosis. We investigated the significance of anti-P. aeruginosa antibodies (precipitins) by crossed immunoelectrophoresis (CIE) in 94 patients attending the Cystic Fibrosis Center of Florence. The highest numbers of precipitins were found in serum from patients chronically colonized in comparison to those patients who were transiently or not colonized. A negative correlation was found between the number of precipitin peaks and clinical conditions, evaluated with Schwachman score, and the number of precipitins and pulmonary functions. In summary, anti-P. aeruginosa antibodies fail to protect against P. aeruginosa bronchopulmonary infections, and are correlated to a more severe disease. Based on our experience, P. aeruginosa antibodies can be considered a reliable index of lung damage.

[Role of viruses in acute gastroenteritis]. / Inquadramento etiopatogenetico delle enteriti acute virali.

In a two years screening carried out on Florentine children we confirmed that Rotaviruses are the most important ethiological agents of acute gastroenteritis in infants less than two years old, irrespective of sex. We had evidence that Rotavirus diarrhea does not occur with the same incidence every year. Rotavirus gastroenteritis is characterized by profuse diarrhea, vomiting, fever and often by respiratory symptoms. Severe complications have not been observed.

Inhibition of viral replication in vitro by antiviral-treated amniotic membrane. Possible use of amniotic membrane as drug-delivering tool.

PURPOSE: To investigate if amniotic membrane (AM) incubated with antivirals can inhibit viral growth in vitro. METHODS: AM samples were incubated with a solution of acyclovir or trifluridine. The treated AM was placed onto monolayers of Vero cells, a continuous cell line from monkey kidney, infected with herpes simplex virus. Viral growth was assessed in comparison to control infected cells by direct examination with an inverted microscope at low magnification for the presence and extension of the typical cytopathic effect, or by estimation of viral genomes. RESULTS: AM soaked in acyclovir or trifluridine inhibited significantly the development of herpes simplex virus in cell cultures, based on the viral growth compared with controls. Non-treated AM did not significantly affect viral replication. CONCLUSIONS: Our preliminary in vitro data show that antiviral-treated amniotic membrane can inhibit viral replication. Therefore, the possibility to combine the previously published anti-inflammatory properties of AM with the capability to absorb antivirals and sustain drug release could be taken into consideration.

Observations on phage-typing of Clostridium difficile: preliminary evaluation of a phage panel.

Of the various methods now employed to type Clostridium difficile, the performance of the phage-typing, as proposed by Sell et al., was investigated on a large collection of isolates from various local epidemiological settings. Because those isolates appeared in preliminary trials mostly resistant to the reference phages, the panel was enlarged by including newly isolated phages. The new set was redundant but better suited to the purpose: out of 287 phage sensitivity patterns 8.4% only were resistant in comparison to the 67.6% resistant to the reference one.

Epidemiological studies of Pseudomonas aeruginosa infections in leg ulcers.

A 6-month prospective study was carried out in an angiology ward on patients with leg ulcers. Pseudomonas aeruginosa was isolated from about half of the samples from the patients. Of the isolates, 85 were serotyped, fingerprinted as to passive pyocin sensitivity and characterized for antibiotic susceptibility pattern. According to all criteria, the isolates belonged to six strains, two of which were clearly epidemic: one infected 11 patients yielding 52 identical isolates, the other infected 4 patients (19 isolates).

In-vitro activity of vancomycin, teicoplanin, daptomycin, ramoplanin, MDL 62873 and other agents against staphylococci, enterococci and Clostridium difficile.

The minimum inhibitory concentrations (MICs) of different antibiotics were determined by a broth microdilution method for staphylococci, enterococci and Clostridium difficile. The antimicrobial agents tested were vancomycin, teicoplanin, daptomycin, ramoplanin, MDL 62873, rifampicin and piperacillin, the latter limited to enterococci. In terms of MIC90S, daptomycin (0.89 mg/l). MDL 62873 (0.99 mg/l), and teicoplanin (1.50 mg/l) were found to be highly active against methicillin-resistant Staphylococcus aureus (MRSA). Daptomycin (MIC90 0.48 mg/l), MDL 62873 (0.95 mg/l) and ramoplanin (1.45 mg/l) were the most active drugs against methicillin-resistant S. epidermidis (MRSE). Teicoplanin (MIC90 0.45 mg/l) was the most active agent against enterococci, followed by MDL 62873 (0.65 mg/l) and daptomycin (1.60 mg/l). MDL 62873 gave the lowest MIC90 (0.17 mg/l) for C. difficile. Teicoplanin (MIC90 0.42 mg/l), daptomycin (0.87 mg/l) and ramoplanin (0.98 mg/l) were also very active. Our results indicate that teicoplanin, daptomycin, ramoplanin and MDL 62873, a teicoplanin derivative, are potentially effective alternative antibiotics for treatment of infections caused by staphylococci, enterococci and C. difficile.