First-time-in-human randomized clinical trial in healthy volunteers and haemodialysis patients with SNF472, a novel inhibitor of vascular calcification.

AIMS: SNF472 is a calcification inhibitor being developed for the treatment of cardiovascular calcification in haemodialysis (HD) and in calciphylaxis patients. This study investigated the safety, tolerability and pharmacokinetics (PK) of intravenous (IV) SNF472 in healthy volunteers (HV) and HD patients. METHODS: This is a first-time-in-human, double-blind, randomized, placebo-controlled Phase I study to assess the safety, tolerability and PK of SNF472 after ascending single IV doses in HV and a single IV dose in HD patients. A pharmacodynamic analysis was performed to assess the capability of IV SNF472 to inhibit hydroxyapatite formation. RESULTS: Twenty HV and eight HD patients were enrolled. The starting dose in HV was 0.5 mg kg-1 and the dose ascended to 12.5 mg kg-1 . The dose selected for HD patients was 9 mg kg-1 . Safety analyses support the safety and tolerability of IV SNF472 in HD patients and HV. Most treatment-emergent adverse events were mild in intensity. No clinically significant effects were observed on vital signs or laboratory tests. PK results were similar in HD patients and HV and indicate a lack of significant dialysability. Pharmacodynamic analyses demonstrated that SNF472 administration reduced hydroxyapatite crystallization potential in HD patients who received IV SNF472 9 mg kg-1 by 80.0 ± 2.4% (mean ± standard error of the mean, 95% CI, 75.3-84.8) compared to placebo (8.7 ± 21.0%, P < 0.001, 95% CI, -32.4 to 49.7). CONCLUSION: The results from this study showed acceptable safety and tolerability, and lack of significant dialysability of IV SNF472. It is a potential novel treatment for cardiovascular calcification in end-stage renal disease and calciphylaxis warranting further human studies.

Effect of antibiotics on biofilm inhibition and induction measured by real-time cell analysis.

AIMS: The purpose of the current work was to study the pattern and dynamics of biofilm formation in clinical isolates of Staphylococcus aureus and Staphylococcus epidermidis in the presence of 10 antibiotics with different action mechanisms. METHODS AND RESULTS: By using impedance measurements in microtitre plates with gold electrodes we have assessed the antibiotic effect on bacterial biofilm growth in real time. The impedance measurements appear to combine both cellular growth and matrix production, representing a measurement of total biofilm mass. Several clinical and reference strains were tested, showing different slopes and cell index values which correlated with their capacity to form biofilms as assessed by attachment to standard microtitre well plates and safranin staining. Biofilms were heavily reduced in biofilm mutants or by protease treatment in protein-based biofilm matrixes. Antibiotic resistance patterns of biofilms, which were very different to those obtained by traditional methods like epsilon-tests on solid media, revealed features that would pass unnoticed by end-point methods. CONCLUSIONS: Once the biofilm is formed, antibiotic efficacy dramatically reduced and sub-inhibitory concentrations of some antibiotics, such as linezolid and clarithromycin, stimulated biofilm growth, stressing the importance of studying antibiotic resistance under biofilm growth conditions in real time. SIGNIFICANCE AND IMPACT OF THE STUDY: Real-time biofilm analysis provides a promising tool to evaluate antibiotic therapy in clinical biofilm-mediated infections.

Effects of chronic hydrogen peroxide exposure on mitochondrial oxidative stress genes, ROS production and lipid peroxidation in HL60 cells.

Hydrogen peroxide (H2O2) is a reactive species that is also involved in the redox regulation of cells because of it is relative stability. In numerous pathological situations, a chronic increase in the production of reactive species is observed, which is related to oxidative stress and cellular damage. This study aimed to evaluate the effects of long-term exposure to different H2O2 concentrations on oxidative stress biomarkers and mitochondrial dynamics in HL60 cells. HL60 cells were treated with a sustained production (0.1, 1.0 and 10.0 nM/s) of H2O2 for one hour. H2O2 production and malondialdehyde (MDA) levels, as a lipid peroxidation marker, increased progressively in HL60 cells in accordance with higher H2O2 exposure, with significant differences between the 10 nM/s H2O2 group and the control and 0.1 nM/s groups. Similarly, progressive increased expression in genes related to the mitochondrial antioxidant defences and mitochondrial dynamics were also observed. Significantly increased gene expression in the 10 nM/s H2O2 with respect to the control group was observed for manganese superoxide dismutase (MnSOD), peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PCG1α), nuclear respiratory factor 2 (Nrf2), mitochondrial transcription factor A (Tfam), mitofusins 1 and 2 (Mfn1 and Mfn2) and uncoupling protein 3 (UCP3), whereas no significant changes were observed in the cytochrome c oxidase subunit IV (COXIV) gene expression. In conclusion, exposure to different sustained production of H2O2 is related to a progressive increase in the gene expression of mitochondrial dynamics and redox processes in HL60 cells, but also to oxidative damage at higher H2O2 production levels.

Prevalence of verotoxigenic Escherichia coli O157 (VTEC O157) and compliance with microbiological safety standards in bovine carcasses from an industrial beef slaughter plant.

UNLABELLED: The presence of Salmonella spp. and levels of Enterobacteriaceae and aerobic plate count were determined in 300 bovine carcasses randomly collected in an industrial cattle slaughterhouse in Catalonia (Spain) as part of a control programme to validate good slaughter practices according to Commission Regulation No 2073/2005. The verotoxigenic Escherichia coli O157 (VTEC O157), although not currently legislated, was also investigated in the same carcasses due to the importance of bovines as a reservoir for this micro-organism. Virulence genes (vtx1, vtx2 and eae), the presence of fliCH 7 and antimicrobial susceptibility were studied in E. coli O157 isolates. Levels of Enterobacteriaceae and aerobic colonies and the presence of Salmonella were within the admissible range stipulated by current legislation. However, VTEC O157 was detected in 14·7% of carcasses. Among the VTEC O157 strains tested for antimicrobial susceptibility, 65% were multiresistant. Overall, the results of this study indicate that even with good manufacturing practices, contamination with VTEC O157 can occur and cattle meat can pose a risk to human health. These results confirm the need for a review of the appropriateness of introducing antimicrobial treatments in the processing of cattle carcasses in Europe. SIGNIFICANCE AND IMPACT OF THE STUDY: This study describes the prevalence of verotoxigenic and multidrug-resistant E. coli O157 strains in bovine carcasses. These results suggest that despite the good manufacturing practices used in the slaughterhouse studied (the largest in Catalonia slaughtering over 81 000 cattle per year), the absence of verotoxigenic E. coli O157 in bovine carcasses cannot be guaranteed.

Scuba diving activates vascular antioxidant system.

The aim was to study the effects of scuba diving immersion on plasma antioxidant defenses, nitric oxide production, endothelin-1 and vascular endothelial growth factor levels. 9 male divers performed an immersion at 50 m depth for a total time of 35 min. Blood samples were obtained before diving at rest, immediately after diving, and 3 h after the diving session. Leukocyte counts, plasma 8oxoHG, malondialdehyde and nitrite levels significantly increased after recovery. Activities of lactate dehydrogenase, creatine kinase, catalase and superoxide significantly increased immediately after diving and these activities remained high after recovery. Plasma myeloperoxidase activity and protein levels and extracellular superoxide dismutase protein levels increased after 3 h. Endothelin-1 concentration significantly decreased after diving and after recovery. Vascular endothelial growth factor concentration significantly increased after diving when compared to pre-diving values, returning to initial values after recovery. Scuba diving at great depth activated the plasma antioxidant system against the oxidative stress induced by elevated pO2 oxygen associated with hyperbaria. The decrease in endothelin-1 levels and the increase in nitric oxide synthesis could be factors that contribute to post-diving vasodilation. Diving increases vascular endothelial growth factor plasma levels which can contribute to the stimulation of tissue resistance to diving-derived oxidative damage.

Oral antiseptics against coronavirus: in-vitro and clinical evidence.

Angiotensin converting enzyme 2 (ACE2) is the cellular receptor for SARS-CoV-2, so ACE2-expressing cells can act as target cells and are susceptible to infection. ACE2 receptors are highly expressed in the oral cavity, so this may be a potential high-risk route for SARS-CoV-2 infection. Furthermore, the virus can be detected in saliva, even before COVID-19 symptoms appear, with the consequent high risk of virus transmission in asymptomatic/presymptomatic patients. Reducing oral viral load could lead to a lower risk of transmission via salivary droplets or aerosols and therefore contribute to the control of the pandemic. Our aim was to evaluate the available evidence testing the in-vitro and in-vivo effects of oral antiseptics to inactivate or eradicate coronaviruses. The criteria used were those described in the PRISMA declaration for performing systematic reviews. An electronic search was conducted in Medline (via PubMed) and in Web of Sciences, using the MeSH terms: 'mouthwash' OR 'oral rinse' OR 'mouth rinse' OR 'povidone iodine' OR 'hydrogen peroxide' OR 'cetylpyridinium chloride' AND 'COVID-19' OR 'SARS-CoV-2' OR 'coronavirus' OR 'SARS' OR 'MERS'. The initial search strategy identified 619 articles on two electronic databases. Seventeen articles were included assessing the virucidal efficacy of oral antiseptics against coronaviruses. In conclusion, there is sufficient in-vitro evidence to support the use of antiseptics to potentially reduce the viral load of SARS-CoV-2 and other coronaviruses. However, in-vivo evidence for most oral antiseptics is limited. Randomized clinical trials with a control group are needed to demonstrate its clinical efficacy.

Outbreak of Legionnaires' disease associated with a supermarket mist machine.

An outbreak of Legionnaires' disease affected 12 customers of a supermarket in a town in Catalonia, Spain, between August and November 2006. An epidemiological and environmental investigation was undertaken. Preliminary investigation showed that all patients had visited the same supermarket in this town where a mist machine was found in the fish section. Water samples were collected from the machine and from the supermarket's water distribution system when high-risk samples were excluded. Environmental samples from the mist machine and clinical samples from two patients tested positive for L. pneumophila serogroup 1 and had the same molecular pattern. The PFGE pattern detected in the clinical and mist-machine isolates had never previously been identified in Catalonia prior to the outbreak and has not been identified since. Four days after turning off the machine, new cases ceased appearing. Molecular study supports the hypothesis that the mist machine from the fish section of the supermarket was the source of infection. We believe it is essential to include exposure to mist machines in any legionellosis epidemiological survey.

[99mTc-MAA peritoneal scintigraphy in pleuroperitoneal comunication in peritoneal dialysis patients]. / Gammagrafía peritoneal con 99mTc-MAA en las comunicaciones pleuroperitoneales en pacientes en diálisis peritoneal.

Peritoneal dialysis is a fully-contrasted alternative for the treatment of end-stage renal disease although it is not exempt of complications. Peritonitis and exit-site infections are among the most frequent complications found. Pleural effusion secondary to pleuroperitoneal communication (PPC) is a serious and uncommon complication in these patients. We present the case of a 50-year old man diagnosed of end-stage renal disease undergoing treatment with peritoneal dialysis who presented progressive dyspnea and right pleural effusion. The peritoneal scintigraphy with (99m)Tc-MAA makes it possible to confirm communication of intraperitoneal dialysis fluid to the pleural cavity.

Simultaneous analysis of saturated and unsaturated oxylipins in 'ex vivo' cultured peripheral blood mononuclear cells and neutrophils.

Oxylipins are a family of saturated and unsaturated fatty acids peroxidation products with bioactive properties. We have developed an improved method for the measurement of ex vivo oxylipin production by peripheral blood mononuclear cells (PBMCs) and neutrophils. We aimed to develop an analytical method to determine the production rates of polyunsaturated fatty acids (PUFAs), PUFA-oxylipin, and saturated-oxylipins by stimulated PBMCs and neutrophils based on solid phase extraction and HPLC-MS/MS technology. A UHPLC system coupled to a Q-Exactive Hybrid Quadrupole-Orbitrap mass spectrometer was used to identify and quantify oxylipin production. For each oxylipin and PUFA their differential response was calculated with respect to a deuterated internal standard factor (ISF). To calculate oxylipin and PUFAs in the culture samples, the individual ISF was used for each oxylipin and PUFA with respect to the deuterated internal standard. PBMCs and neutrophils showed a different pattern of oxylipin production and fatty acid secretion. Lipopolysaccharide (LPS) did not stimulate oxylipin production or fatty acids secretion in PBMCs, whereas phorbol myristate acetate (PMA) stimulation increased the production rate of 5-HETE, 15-HETE, 15-HEPE, 17-DoHE, PGE2, AA, and DHA. LPS stimulation decreased 16-hydroxyl-palmitatte (16-OHPAL) production and DHA secretion in neutrophils, while PMA stimulation increased the production rate of AA and its derivate oxylipins, 5-HETE, 15-HETE, and PGE2. In conclusion, we have developed a new method to determine oxylipins derived from both saturated and unsaturated fatty acids in culture cell media. This method has enough sensitivity, and accuracy, to determine oxylipin production and fatty acid secretion by PBMCs and neutrophils.

Effects of exercise intensity on lymphocyte H2O2 production and antioxidant defences in soccer players.

OBJECTIVE: Physical exercise is capable of enhancing or suppressing the immune response depending on the intensity and duration of exercise. This study investigated how exercise intensity influences the lymphocyte antioxidant response and the induction of cellular oxidative damage. DESIGN: Eighteen voluntary male pre-professional soccer players participated in this study. Sportsmen played a 60 min training match, and were divided into three groups depending on the intensity degree during the match: low, medium and high intensities. MEASUREMENTS: Malondialdehyde (MDA), vitamins C and E and haem oxygenase-1 (HO-1) gene expression were measured in lymphocytes. Reactive oxygen species (ROS) production was determined in lymphocytes and neutrophils. RESULTS: Lymphocyte MDA levels and H(2)O(2) production were significantly increased in the group which performed the most intense exercise. Neutrophil counts and ROS production increased progressively with the exercise intensity. Vitamin C significantly decreased after exercise in the highest-intensity group in comparison with initial values, whereas vitamin E levels significantly increased in the medium and high-intensity groups. HO-1 gene expression significantly increased in the medium and high-intensity groups. CONCLUSIONS: Exercise intensity affects the lymphocyte and neutrophil oxidant/antioxidant balance, but only exercise of high intensity induces lymphocyte oxidative damage.

SNF472, a novel anti-crystallization agent, inhibits induced calcification in an in vitro model of human aortic valve calcification.

The purpose of the present study was to investigate whether SNF472, the hexasodium salt of myo-inositol hexaphosphate (IP6 or phytate): 1. Inhibits induced calcification in cultured aortic valve interstitial cells (VIC) as an in vitro model of aortic valve stenosis and 2. Whether inhibition is different in VIC obtained from healthy and calcified aortic valves. VIC from healthy (n = 5) and calcified (n = 7) human aortic valves were seeded in basic growth medium, osteogenic differentiation medium alone, or in osteogenic medium with SNF472 (3, 10, and 30 µM) and cultivated for 3 weeks. Calcification was quantified spectrophotometrically after Alizarin Red staining. In VIC from calcified valves, a complete inhibition of calcification was observed with SNF472 concentrations of 10 and 30 µM (p < .01), significantly stronger than in VIC from healthy valves. When SNF472 was added to VIC after 1 week in osteogenic medium, 30 and 100 µM SNF472 inhibited the progression of ongoing calcification by 81 and 100% (p < .01), respectively. The same concentrations of SNF472 given after 2 weeks reduced calcification by 35 and 40% respectively (not significant). SNF472 inhibited both the formation and the progression of calcification with the strongest effect in VIC from calcified valves.

[Utility of myocardial perfusion SPECT for evaluation of patients from Chest Pain Unit]. / Análisis y utilidad de la tomografía de emisión de fotón único de perfusión miocárdica en la evaluación de pacientes de la Unidad de dolor torácico.

UNLABELLED: The Chest Pain Units (CPU) are currently the best solution to improve management of patients with acute chest pain in the Emergency Room thanks to the use of reliable ischemia diagnostic detection tests and early treatment. OBJECTIVE: To assess the value of myocardial perfusion SPECT (MPS) in the CPU in order to treat acute coronary syndromes (ACS) early and discharge patients with low risk of coronary artery disease (CAD) who can be treated as outpatients. MATERIAL AND METHODS: We studied 629 patients from January 2003 to September 2005 with acute chest pain suggestive of angina, normal cardiac enzymes and normal or non-diagnostic ECG who had been referred to Nuclear Medicine for evaluation with a stress test for ischemia: 32 p treadmill stress testing and 597 p MPS (525 p exercise-rest and 72 p pharmacologic stress test). We compared the results with catheterization and clinical follow up for a 6-months period, evaluating new coronary events. RESULTS: 76 % of MPS were normal and 24 % pathological. Only 1.5 % of the patients with normal MPS had CAD or coronary events in the follow-up, increasing to 35.2 % in patients with ischemia. A total of 45 catheterizations were performed, showing CAD 27 (24 with pathological MPS). A total of 2.6 % of the patients had coronary events during follow-up, 75 % of whom had pathological MPS. CONCLUSION: The MPS improves diagnosis of ACS in the CPU, with a very low number of new coronary events at 6 months of the follow-up and permits safe discharge of these patients.

SNF472, a novel inhibitor of vascular calcification, could be administered during hemodialysis to attain potentially therapeutic phytate levels.

BACKGROUND: Cardiovascular calcification (CVC) is a major concern in hemodialysis (HD) and the loss of endogenous modulators of calcification seems involved in the process. Phytate is an endogenous crystallization inhibitor and its low molecular mass and high water solubility make it potentially dialyzable. SNF472 (the hexasodium salt of phytate) is being developed for the treatment of calciphylaxis and CVC in HD patients. We aimed to verify if phytate is lost during dialysis, and evaluate SNF472's behaviour during dialysis. METHODS: Dialyzability was assessed in vitro using online-hemodiafiltration and high-flux HD systems in blood and saline. SNF472 was infused for 20 min and quantified at different time points. RESULTS: Phytate completely dialyzed in 1 h at low concentrations (10 mg/l) but not when added at 30 or 66.67 mg/l SNF472. In bypass conditions, calcium was slightly chelated during SNF472 infusion but when the system was switched to dialysis mode the calcium in the bath compensated this chelation. CONCLUSION: Phytate dialyses with a low clearance. The administration of SNF472 as an exogenous source of phytate allows to attain supra-physiological levels required for its potential therapeutic properties. As SNF472 is infused during the whole dialysis session, the low clearance would not affect the drug's systemic exposure.

Scuba diving enhances endogenous antioxidant defenses in lymphocytes and neutrophils.

The aim was to study the effects of a scuba diving session on the lymphocyte antioxidant system, NO synthesis, the capability to produce reactive oxygen species and the antioxidant response in neutrophils. For that purpose seven male divers performed an immersion at a depth of 40 m for 25 min. The same parameters were measured after an hyperbaric oxygen (HBO) treatment at resting conditions in a hyperbaric chamber. Lymphocyte H2O2 production rose after diving and after HBO treatment. Glutathione peroxidase (GPx) and catalase activities increased after diving in lymphocytes, while after HBO exposure only increased GPx activity. Lymphocyte HO-1 mRNA expression increased after diving and after HBO exposure, while iNOS levels and nitrite levels significantly increased after diving. The hyperoxia associated to scuba diving leads to a condition of oxidative stress with increased lymphocyte H2O2 production, HO-1 expression, NO synthesis and antioxidant enzyme adaptations in order to avoid oxidative damage.

A novel pharmacodynamic assay to evaluate the effects of crystallization inhibitors on calcium phosphate crystallization in human plasma.

Cardiovascular calcification (CVC) is a progressive complication of chronic kidney disease and a predictor of CV events and mortality. The use of biomarkers to predict CV risk and activities of potential or current treatment drugs in these patients could have a crucial impact on therapeutic approaches. Our aim was to develop a novel assay for measurement of the rate of calcium phosphate crystallization in human plasma and provide a tool to evaluate the effects of crystallization inhibitors. The efficacy of inhibitors was determined by adding inhibitory compounds (polyphosphates, fetuin-A, sodium thiosulfate or citrate) to control samples. The assay was additionally validated for SNF472, an experimental formulation of phytate being developed for the treatment of calciphylaxis and CVC in patients with end-stage renal disease (ESRD) undergoing hemodialysis (HD). The method was repeatable and reproducible. The plasma crystallization rate was reduced up to 80% in a concentration-dependent manner following treatment with inhibitors in vitro, among which SNF472 was the most potent. This method appears beneficial in evaluating and discriminating between inhibitory activities of compounds such as polyphosphates on calcium phosphate crystallization, which present a novel therapeutic approach to treat CVC in ESRD patients.

Vitamin C supplementation influences the antioxidant response and nitric oxide handling of erythrocytes and lymphocytes to diving apnea.

OBJECTIVE: We have investigated the influence of vitamin C diet supplementation on the antioxidant response and nitrite levels in lymphocytes and erythrocytes during diving apnea. SUBJECTS: Seven male professional apnea divers participated in a double blind crossover study. Divers were randomly assigned to either vitamin C supplemented or placebo groups. The subjects did not take any other supplements than the ones provided for this study. INTERVENTION: One group was supplemented with vitamin C capsules (1 g per day) for 7 days while the other group took a placebo composed of lactose. The usual dietary habits of participants were assessed using a self-reported 7-days 24-h recall before the day of the study. Blood samples were taken under basal conditions, immediately after diving apnea for 4 h and after 1 h of recovery. RESULTS: Catalase activity increased in erythrocytes (23%) and superoxide dismutase increased in lymphocytes (35%) during the recovery only in the placebo group. Lymphocyte ascorbate levels increased in the supplemented group after diving (85%) and maintained high at recovery. Plasma nitrite levels increased about twofold in both groups during the recovery. Erythrocyte nitrite levels increased after diving (50%) and about twofold during the recovery in the supplemented group. Nitrite levels and iNOS levels in lymphocytes were higher in the placebo group than in the supplemented during the recovery. Erythrocyte carbonyl derivates were unchanged in all situations. CONCLUSIONS: Vitamin C supplementation influenced the antioxidant response and NO handling in erythrocytes and lymphocytes to the oxidative stress induced by hypoxia-reoxygenation.

[Iatrogenic lung microembolism detected by 18FDG PET/CT]. / Microembolia pulmonar iatrogénica detectada mediante 18F-FDG PET/TC.

We present the cases of two oncology patients: a male with Hodgkin's disease after completion of chemotherapy, and a woman recently diagnosed of melanoma, who underwent positron emission tomography/computed tomography (PET/CT) with 18F-FDG for therapeutic monitoring and initial staging, respectively. In both cases, hypermetabolic foci of 18F-FDG in lung parenchyma were found, without morphologic abnormalities in CT. These findings would have been consistent with lung pathology in the absence of any anatomic correlation. Combined PET/CT interpretation was of lung microembolisms probably originated at the injection site.

In vitro activities of 22 antimicrobial agents against Listeria monocytogenes strains isolated in Barcelona, Spain. The Collaborative Study Group of Listeriosis of Barcelona.

The in vitro activity of 22 antimicrobial agents against 82 human Listeria monocytogenes strains isolated in Barcelona from 1994 to 1998 was determined. Ampicillin and gentamicin showed good in vitro activity against all strains (MIC90: 1 and < or = 0.25 microg/ml, respectively). No resistance to rifampin or co-trimoxazole was detected and only one strain was resistant to tetracycline. Of the nine fluoroquinolones tested, clinafloxacin and gemifloxacin were the most active compounds (MIC90: 0.12 and 0.25 microg/ml, respectively). No increasing MICs values were observed during the five-year period.

Initial numbers, serovars and phagevars of Listeria monocytogenes isolated in prepared foods in the city of Barcelona (Spain).

A total of 1100 samples of prepared foods purchased in restaurants and delicatessen shops in the city of Barcelona was examined for the presence of Listerio. L. monocytogenes was more frequently isolated from foods intended to be consumed without further cooking (9.3%) than from foods intended to receive further cooking prior to consumption (2.9%). A quantitative study, carried out with 773 samples, yielded 1% of samples with numbers of L. monocytogenes higher than 100 CFU/g. Strains of L. monocytogenes belonged to serovar 4b (36.3%), 1/2a (29.5%), 1/2b (22.7%) and 1/2c (11.3%). L. monocytogenes serovar 4b phagevar 3274: 2671 (6 of 11) was prevalent among the strains studied.

Incidence of Listeria monocytogenes in fresh foods in Barcelona (Spain).

From September 1989 to March 1990, a study of Listeria spp. was carried out on 311 samples of raw foodstuffs from markets and other establishments in the city of Barcelona. These foodstuffs included vegetables (103 samples), minced meats from pork, beef and poultry (168 samples) and bivalve molluscs (40 samples). L. monocytogenes was isolated in 7.8% of the vegetable samples, 17.3% of the minced meats and in 7.5% of bivalve molluscs. The most frequent serovars were 1/2 and 4. Other species isolated were L. innocua, L. welschimeri and L. seeligeri.