Prevalence, serotypes and virulence genes of Shiga toxin-producing Escherichia coli isolated from ovine and caprine milk and other dairy products in Spain.

The aim of this study was to determinate the prevalence, serotypes and virulence genes of Shiga toxin-producing Escherichia coli (STEC) strains isolated from different dairy products (DP) in Spain with the purpose of determining whether DP represent a potential source of STEC pathogenic for humans. A total of 502 DP were examined from 64 different ovine and caprine flocks and 6 dairy plants in Extremadura (Western Spain). Samples were collected monthly between March 2003 and June 2004 and included 360 unpasteurised milk obtained from the bulk tank, 103 fresh cheese curds and 39 cheeses. Samples obtained were examined for STEC using genotypic (PCR) methods. STEC strains were detected from 39 (10.8%) bulk tank, 4 (3.9%) fresh cheese curds and 2 (5%) cheese, whereas O157:H7 serotype were isolated from one (0.3%) bulk tank. A total of 9 STEC strains (O27:H18, O45:H38, O76:H19, O91:H28, O157:H7, ONT:H7, ONT:H9 and ONT:H21) were identified in this study. One of them, the serotype O27:H18, has not been reported previously as STEC. PCR showed that 3 strains carried stx1 genes, 5 possessed stx2 genes and 1 both stx1 and stx2. Whereas all STEC caprine isolates showed ehxA genes, only O157:H7 serotype showed eae virulence genes. The strain O157:H7 isolated possessed intimin type gamma1 and belonged to phage type 31. This study confirms that dairy product is an important reservoir of STEC pathogenic for humans.

Bovine tuberculosis in wild boar (Sus scrofa), red deer (Cervus elaphus) and cattle (Bos taurus) in a Mediterranean ecosystem (1992-2004).

During the last 12 years, an increasing frequency in condemnation of hunted red deer and wild boar carcasses due to the presence of tubercle-like lesions has been observed in Extremadura (Western Spain). Before 1993, tuberculosis was a very rare finding in hunted animals. The current tuberculosis regional prevalence in cattle approaches 0.4% after years of expensive test and slaughter campaigns. It is imperative to investigate the epidemiology of Mycobacterium bovis infection in red deer and wild boar in order to keep a good health status and to maintain the effectiveness of domestic species TB eradication programs. The present paper evaluates the problem in Sierra de San Pedro, estimating the prevalence of TB in wild boar and red deer, the main wild artiodactyls in the area, and domestic cattle since 1992-2004, by the use of a low-cost surveillance method based on detailed pathological inspection of hunted animal carcasses. Microbiology and molecular epidemiology studies on several M. bovis isolates from domestic and wild animals helped to define the interspecies contacts. These findings, as well as recent history of game estates management and descriptive epidemiology field work, throw light on the rise and maintenance of these epizootics.

An epidemiological evaluation of Mycobacterium bovis infections in wild game animals of the Spanish Mediterranean ecosystem.

Recreational hunting of indigenous wild artiodactyls has been one of the most lucrative and rapidly growing industries in Western Spain over the last five years. In the absence of careful ecological management, one consequence of the commercial exploitation of this natural resource has been the appearance of outbreaks of infectious disease; most notably bovine tuberculosis. From the outset of the study in 1997, we have observed a steady increase in prevalence of Mycobacterium bovis (M. bovis) in both species reaching 1.74 (+/-0.17) in deer in 2002 and 2.32 (+/-0.24) in wild boar. The latter species seems to be most severely affected with pulmonary lesions appearing more chronic than those observed in deer. In this study, we describe the epidemiology of M. bovis in European wild boar (Sus scrofa) and Iberian red deer (Cervus elaphus hispanicus) in Extremadura (W. Spain); a region where there are large areas of natural habitat for these species.

Estudio comparativo del nivel hepático de metales pesados y metaloides en aves rapaces diurnas de Galicia y Extremadura / Comparative study of heavy metal and metalloid hepatic levels in diurnal raptors from Galicia and Extremadura

Las aves rapaces son predadores situados en la cumbre de las cadenas alimenticias, por lo cual podrían ser consideradas como adecuados bioindicadores en los programas de biomonitorización del medio, a fin de evaluar la presencia y efecto de diversos contaminantes en los ecosistemas. En el presente trabajo se han determinado las concentraciones de diversos metales pesados y metaloides (As, Cd, Pb y Zn) en el tejido hepático de distintas especies de aves rapaces diurnas procedentes de Galicia y Extremadura, empleando para ello animales que llegaron muertos o se sacrificaron a su llegada a los Centros de Recuperación de Fauna Salvaje de ambas comunidades. Tras la digestión de las muestras por vía húmeda, el contenido de elementos inorgánicos fue determinado por medio de Espectrometría de Masas con fuente de plasma a copl ado por inducción (ICP-MS). L as concentraciones cuantificadas de los cuatro elementos se situaron en general dentro de los niveles considerados como normales para las aves, no causando efectos patológicos directos. Destaca el hecho de que de los cuatro elementos considerados , las concentraciones de Cd y Zn cuantificadas en Galicia fueron siempre ligeramente superiores a las obtenidas en Extremadura. Por otra parte, considerando la especie a estudio, destaca el hecho de que los valores más elevados de los cuatro elementos analizados se correspondieran con ejemplares de Busardo ratonero procedentes de Galicia (AU)

Raptors are predators located at the top of food chains, which offers the possibility of use these species as adequate bioindicators within the environmental biomonitoring programs, in order to assess the presence and effect of a broad spectrum of contaminants on the ecosystems. In the present work, heavy metal and metalloid (As, Cd, Pb and Zn) contents in liver of different raptor species from Galicia and Extremadura have been determined, using animals that arrived died or that were sacrificed after their arrival to the Wildlife Recovery Centres of both communities. After wet digestion of the sample, inorganic element content was determined using inductively coupled plasma- mass spectrometry (ICP-MS). The concentrations of all the analyzed elements were in general close to those considered as normal in avian species, and therefore do not produce direct pathological effects. It must be emphasized that the hepatic concentrations of Cd and Zn quantified in Galicia were slightly higher than those corresponding to Extremadura. Moreover, and when considering the studied species, it must be signalled that the highest values for the four analyzed elements were observed in Common buzzard from Galicia (AU)

Manejo de osteoporosis en atención primaria / Osteoporosis management in primary care

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Isolation of Cryptococcus species including C. neoformans from cloaca of pigeons.

Upper digestive tract of the pigeon (Columba livia) is well known as a reservoir for different species of Cryptococcus, but lower portions are not so frequently studied. In the present study, we tested on selective media a total of 331 pigeon cloacal swabs; Cryptococcus spp. were recovered from 26 (7.85%). Cryptococcus uniguttulatus was isolated from 11 samples (3.32%), C. laurentii from six (1.81%), C. neoformans var. neoformans from six (1.81%) and C. albidus from three of them (0.91%). The results show the importance of pigeon in the cryptococcosis epidemiology as reservoir and carrier for C. neoformans var. neoformans, but also for other Cryptococcus species of increasing clinical interest.

Epidemiology of Mycobacterium bovis infections of pigs and wild boars using a molecular approach.

A molecular epidemiological approach was applied to establishing a possible role for the wild boar as a natural reservoir of Mycobacterium bovis in Sierra de Villuercas, Western Spain; an area free of farmed cattle and wild deer populations. Spoligo and VNTR typing were used over a three year period to study the epidemiological relationship between the occurrence of bovine tuberculosis (TB) in extensively bred Iberian pigs and indigenous wild boar. The 37 sampled wild boar showed different degree of calcified granulomatous lesions in retropharyngeal, mediastinal and pulmonary lymph nodes. The 25 sampled Iberian pigs showed calcified lesions, mainly in the respiratory tract. Lesions located in the mesenteric lymph nodes appeared secondarily. M. bovis was isolated from all affected animals. Twenty-five and 37 isolates of M. bovis were obtained from domestic pigs and wild boar, respectively. Our findings provide evidence that supports the possibility of cross infection between wild boar and domestic pig populations. This is contrary to the generally held belief that swine represent an epidemiological dead end host and play no role in the epidemiology of M. bovis.

[Classification of osteoporosis. Risk factors. Clinical manifestations and differential diagnosis]. / Clasificación de la osteoporosis. Factores de riesgo. Clínica y diagnóstico.

The fact that there are multiple factors that can intervene in the ethiopathology of osteoporosis, together with its scarce clinical expression until complications -fractures- are produced, poses a practical problem in medical practice when it comes to identifying those persons at risk of osteoporosis. These aspects, together with the difficulties in obtaining access to a complementary test that would confirm the diagnosis and the overload of tasks in our clinic, mean that the disease often passes unnoticed. Hence, in the section on diagnostic orientation of osteoporosis we attempt to clarify and give cohesion to this approach, offering highly useful guidelines for suspecting the presence of the disease and patterns of action that will make it possible to diagnose in a more efficient and accurate way

[Optimization of the conditions for RAPD-PCR of Candida spp. and Cryptococcus spp]. / Optimización de las condiciones RAPD-PCR en Candida spp. y Cryptococcus spp.

In this study, we determined the optimal RAPD amplification conditions to obtain genetic molecular markers for the rapid and accurate identification of Cryptococcus spp. and Candida spp. The following parameters are modified: template DNA, DNA polymerase, magnesium cloride and primer concentration; denaturation, annealing and extension time, temperature of annealing and thermal cycles. After the optimization, reliable and reproducible RAPD patterns are obtained.

[Epidemiological study of cryptococcosis in Spain: first results]. / Estudio epidemiologico de la criptococosis en España: primeros resultados.

The study constitutes an approach to the knowledge of the epidemiology of cryptococosis in Spain. For detection of cases 167 Spanish hospitals were contacted. All cases included were accompanied by the correspondent isolate of Cryptococcus neoformans, together with clinical, demographic and mycological data. Results obtained from January 1998 to end of December 1999 are analysed and presented here. Fifty-six Spanish hospitals reported 58 cases of cryptococcosis; only 43 of them were adequately documented and accompanied by the clinical isolate. The results showed a higher incidence in males (88.4%) than in females (11.6%); being most frequently affected those between 30 and 40 years old (48.8%). The 84.6% (33) corresponded to new cases and 15.4% (6) to relapses of the disease. The HIV infection was the most frequent risk factor reported (86%) and, for 29.7% (11) of them, cryptococcosis was the AIDS defining disease. For the diagnosis, CSF analysis showed the best results (India ink; culture and antigen detection). All strains collected (100%) corresponded to C. neoformans variety neoformans. Serotypes distribution was 45.5% for serotype A and 22.7% for each of serotypes D and AD.

Mastitis caused by Mycoplasma mycoides subspecies mycoides (large colony type) in goat flocks in Spain.

We describe three different outbreaks of mastitis caused by M. mycoides subspecies mycoides LC type (Mmm LC) in three goat flocks from the Extremadura Region of south-west Spain. Thirty-two fast-growing isolates were obtained on Hayflick's and Friis's media with inhibitors from different specimens. All were identified as Mmm LC in spite of their cultural, biochemical and serological features.

Rapid and easy method to extract and preserve DNA from Cryptococcus neoformans and other pathogenic yeasts.

The mucopolysaccharide capsule of Cryptococcus neoformans and other pathogenic yeasts prevent the extraction of DNA from these important zoonotic agents. We report that the use of a lysis buffer containing a high concentration of urea is an easy, efficient and time-saving technique to obtain high yields of good-quality DNA for molecular diagnosis. The use of urea also prevents the degradation of DNA during storage of samples at room temperature for up to 6 months.

An indirect fluorescent antibody technique for detection of anti-Dermatophilus congolensis antibodies in sheep.

An indirect fluorescent antibody (IFA) technique has been developed for detection of anti-Dermatophilus antibodies in sheep. Sera from 25 bacteriologically confirmed clinically affected sheep and from 10 negative non affected lambs were used. Whole cell antigen from brain heart infusion cultures of D. congolensis was used and all sera were tested in the same way for cross-reactivity against antigens obtained from cultures of Actinomyces viscosus, Micrococcus luteus, Nocardia asteroides, and Corynebacterium pseudotuberculosis. Sera from Dermatophilus-infected sheep gave positive results with D. congolensis antigen and negative results with the antigens from other bacteria. The whole cell antigens employed were simple to prepare and easy to recognise by microscopy. Cross-reactivity was further tested using the D. congolensis culture whole cell antigen and 3 sera from sheep with bacteriologically confirmed natural infections due to Corynebacterium pseudotuberculosis, Actinomyces pyogenes and Nocardia asteroides. None of these sera showed positive reactions. The authors recommend this technique for serological surveys and research on dermatophilosis.

In vitro studies of Dermatophilus congolensis antimicrobial susceptibility by determining minimal inhibitory and bacteriocidal concentrations.

The Minimal Inhibitory Concentration (MIC) and Minimal Bacteriocidal Concentration (MBC) of 19 antimicrobials on 16 isolates of D. congolensis were determined. The potential field efficacy of the agents was evaluated by comparing the results with serum levels of drug unbound to proteins and the in vitro and in vivo findings of other authors. A modified standard microtechnique was used for serial dilution-antimicrobial sensitivity and found to be easy and reproducible. Erythromycin, spiramycin, penicillin G, ampicillin, chloramphenicol, the streptomycin, amoxicillin, the tetracyclines and novobiocin had high serum concentrations in comparison with their MBCs and were shown to have potential use for the treatment of dermatophilosis.

Enzymatic activities of Dermatophilus congolensis measured by API ZYM.

API ZYM kit was used to test enzymatic activities on eighteen strains of Dermatophilus congolensis. All strains produced lipase and acid phosphatase, which act on lipids, and leucine arylamidase which act on proteins. Another 10 exoenzymes were present in at least one of the strains.