Alfonso Maldonado

Risk factors associated with the antimicrobial resistance of staphylococci in canine pyoderma

This study reports the susceptibility to antimicrobial agents of staphylococci (n=105) isolated from dogs, and the factors associated with this resistance. The study animals were 23 healthy dogs (group A), 24 with first-time pyoderma (group B), and 27 with recurrent pyoderma that had undergone long-term antibiotic treatment (group C). Staphylococci were more commonly isolated from the pyoderma-affected than the healthy dogs (p<0.0001). Some 78% of the isolates were resistant to at least one antimicrobial agent. Resistance to amoxicillin-clavulanate, cephalosporins (OR 4.29, 95% CI [1.15, 16.3] respectively), enrofloxacin (OR 9.47, 95% CI [1.53, 58.5]) and ciprofloxacin (OR 79.7 95% CI [3.26, 1947.4]) was more common among group C isolates. Some 32% of all the isolates were multiresistant (MR) and 10.4% were methicillin-resistant (MRS). The probability of isolating MRS staphylococci in group C increased by a factor of four (95% CI [1.18, 17.9]) compared to A plus B. Multi-resistant (MR) isolates were obtained more commonly from urban than rural dogs (OR 3.79, 95% CI [1.09, 13.17]). All the MRS staphylococci encountered were obtained from urban dogs and more commonly from male dogs (p=0.07). This study shows that dogs bred in urban habitat, with a history of antibiotic therapy in the past year represents significant risk of being carriers of isolates resistant to methicillin (MRS) and other antimicrobials. These factors should be considered before applying an antimicrobial treatment in veterinary clinics.

An evaluation of the application of infrared thermal imaging to the tele-diagnosis of sarcoptic mange in the Spanish ibex (Capra pyrenaica).

The application of infrared thermal imaging to the diagnosis of sarcoptic mange in the wild Spanish ibex (Capra pyrenaica) was evaluated. Seventy-three ibexes (35 males, 38 females) of varying ages were studied. Each animal was observed using conventional binoculars (OT) to detect lesions characteristic of mange. Infrared thermography (IR) was then performed and the resultant image judged negative or positive. The distance from the thermograph to the animal was measured, and the animal killed. Skin samples were taken for mite detection by routine laboratory diagnosis (LAB). The most sensitive and specific technique for the tele-diagnosis of sarcoptic mange in the Spanish ibex is OT, as it permits diagnosis over greater distances than IR, which sensitivity is impaired at distances >100m. When disease prevalence is low, such as in initial and final phases of an epidemic, a more sensitive technique would be valuable in detecting all affected animals.

Salmonella prevalence and characterization in a free-range pig processing plant: Tracking in trucks, lairage, slaughter line and quartering

New consumer tendencies are focused on products derived from systems which allow both a high animal welfare condition and a high food safety level. However, sometimes animal welfare regulations make the adoption of adequate bio-security measures difficult, representing a barrier for animal health and food safety. Thus the aim of this study was to determine the prevalence of Salmonella at different points of the pig slaughtering process (Trucks, Lairage, Slaughter line and Quartering, TLSQ) from pigs reared in free-range systems. From eight samplings a total of 126 Salmonella isolates out of 1160 different samples were recovered (10.86%). The highest percentage of isolates was detected at the points of pre-scalding (29/80, 36.25%), trucks (13/56, 23.21%), cecal contents (17/80, 21.25%), tonsils (14/80, 17.50%), ileocecal lymph nodes (13/80, 16.25%) and lairage (9/64, 14.06%). Furthermore, eighteen isolates were obtained from different environmental samples from slaughter line and quartering plant (knives and surface of tables) (5.63%) and three isolates at the quartering plant samples (ham, shoulder and loin) (3.75%). Fourteen different serotypes were isolated: Bredeney, Rissen, Derby, Typhimurium, Montevideo, Israel, Anatum, Emek, Monophasic Salmonella Typhimurium (mST), Choleraesuis, Durban, Kentucky, London and Sandiego. S. Typhimurium phage types U311, 193, 104b and UT were identified. Moreover, mST strain was phage typed as U311. From TLSQ1, TLSQ2 and TLSQ4, different strains of S. Derby, S. Rissen and S. Bredeney serotypes were isolated from pig and environmental samples, pointing to a potential cross contamination. Molecular typing (Pulsed-Field Gel Electrophoresis, PFGE) of these strains confirmed the cross contamination. In the remaining samplings, different serotypes were obtained in each sampled point of the chain, assuming that the isolated serotypes belonged to different epidemiological origins. Our results show the isolation of different serotypes of Salmonella spp. from both pigs and environmental samples, which constitutes a great risk for the contamination of pork from free-range pigs both prior and post slaughter. These data support the intensification of the cleaning and disinfection in the pre-slaughter environment (i.e. trucks, lairage), especially when a higher workload is present, as well as the inclusion of new strategies to decrease or eliminate the risk of Salmonella spp. infection or recontamination from the environment in pork from organic or eco-friendly systems.

Streptococcus suis serotypes associated with different disease conditions in pigs

the yeast in the oral cavity whereas selenium sulphide had no effect (Table 2). Similarly, the frequency of isolation ofM pachydermatis from the oral cavity of the miconazole-chlorhexidine treated dogs (seven of 15) was significantly reduced (P<0.05), whereas selenium sulphide treatment had no such effect (M pachydermatis isolated in 12 of 17 dogs). This study indicates that effective treatment of M pachydermatisassociated seborrhoeic dermatitis with miconazole-chlorhexidine shampoo is not only associated with reductions ofM pachydermatis populations on the skin surface but also in the oral cavity. This might indicate that oral carriage reflects contamination of the mucosae by constant seeding from a reservoir of the yeast on heavily colonised skin, particularly in pruritic dogs. Alternatively, reduced yeast counts in the mouth could have resulted from a direct antifungal effect by transfer of the shampoo to the oral mucosae by licking. Regardless of the mechanism, the reduction in oral cavity populations may be of therapeutic value in reducing transfer of oral populations back to the skin. It is noteworthy that only the more effective of the two shampoos studied had a significant effect on oral carriage. Further studies are required to determine whether there is significant transfer ofM pachydermatis cells between skin and mucosal sites in dogs, and to determine whether treatment of mucosal carriage sites may be of value in the management of recurrent cases of dermatitis in a manner similar to that recently suggested for canine pyoderma (Saijonmaa-Koulumies and others 1998).

Suilysin production by Streptococcus suis strains isolated from diseased and healthy carrier pigs in Spain

I. Luque, DVM, Departamento Patologia Infecciosa, Edificio Sanidad Animal, Campus Universitario de Rabanales, Ctra de Madrid, s/n, 14071C6rdoba, Spain SINCE Streptococcus suis was first reported as a cause of disease in pigs in the Netherlands and the UK, new cases have been cited in major swine-producing countries around the world (Windsor and Elliot 1975, Sanford and Higgins 1992). S suis is responsible for a variety of pathological conditions, such as septicaemia, meningitis, arthritis, bronchopneumonia, and reproductive failures in pigs, with important losses in production in herds (Sanford and Higgins 1992, Luque and others 1998a). It is now generally accepted that among strains within one serotype of S suis there are differences in virulence (Vecht and others 1991, Jacobs and others 1994, Gottschalk and others 1998). Different bacterial structures or products have been considered to be virulence factors (Jacobs and others 1994, Salasia and Lammler 1994). Vecht and others (1991) described two proteins, a muramidase-released protein (MRP), which is a 136 kD cell wall-associated protein, and an extracellular factor (EF), a 110 kD protein, both related to the virulence of the strains in pigs. Jacobs and others (1994) described a thiol-activated haemolysin called suilysin, a secreted protein, which can also be a virulence factor of S suis. The aim of this study was to determine the production of suilysin by studying both strains isolated from diseased pigs (clinical strains) and strains isolated from healthy carrier pigs (tonsillar strains) in Spain, showing any possible relation with other virulence-related proteins (MRP and EF proteins) which had already been studied (Luque and others 1998b). A total of 203 swine strains were studied, 85 of them isolated from pigs with different clinical conditions (septicaemia, meningitis, arthritis, reproductive failure, bronchopneumonia, abscess formation) and the remaining 118 isolated from healthy carrier pigs. Several processes such as isolation, biochemical and serological identification, and production ofMRP and EF proteins, were carried out as described by Luque and others (1998b). Production of suilysin was carried out by using a microtitre assay as described by Jacobs and others ( 1994). Colonies from Columbia agar base (Oxoid) were inoculated in 100 ml Todd-

Septicaemic pasteurellosis in free-range pigs associated with an unusual biovar 13 of Pasteurella multocida

Biochemical profiles, PFGE typing and MLST analysis were used to investigate an outbreak of septicaemic pasteurellosis in a free-range pig farm in Spain. Signs of coughing, dyspnoea and a visible inflammation of the ventral area of the neck (jowl), which acquired a cyanotic and necrotic appearance, were the characteristic findings in affected animals, associated with a high morbidity (70%) and case mortality (95%). Diffuse, haemorrhagic and fibrinous pleuroneumonia and acute, focally extensive and haemorrhagic myositis and panniculitis were observed in the histopathological analysis from three analyzed animals. Pasteurella multocida subsp. multocida, capsular type B, biovar 13 was isolated in pure culture from lung, submandibular tissue (jowl), liver, spleen and kidney tissue from diseased pigs. After PFGE typing, all P. multocida isolates displayed undistinguishable macrorestriction patterns with Bsp120I restriction enzyme demonstrating that the infection was caused by a single strain. With the multihost P. multocida MLST database, all P. multocida isolates were assigned to the new sequence type ST47 which was highly related with other bovine isolates of P. multocida type B associated with haemorrhagic septicaemia. This is the first description of an outbreak of septicaemic pasteurellosis in free-range pigs associated with P. multocida type B of the unusual biovar 13. The communication and complete diagnosis of cases of swine septicaemia and the possible role of pigs as reservoirs of this new pathogen must be evaluated to determine the importance of this disease for pigs.

Characterization of the immune response and evaluation of the protective capacity of rSsnA against Streptococcus suis infection in pigs

The efforts made to develop vaccines against Streptococcus suis have failed because of lack of common antigens cross-reactive against different serotypes of this species. The cell wall-anchored proteins can be good vaccine candidates due to their high expression and accessibility to antibodies, among these, a cell-wall protein, DNA-nuclease (SsnA), present in most of the S. suis serotypes and clinical isolates collected from infected pigs, was selected. An experimental challenge against S. suis serotype 2 in a pig model was used to validate the efficacy of recombinant SsnA combined with aluminium hydroxide plus Quil A as adjuvants, previously tested in mice by our research group with good results. In our study, clinical characteristics, bacterial load and spread, haematological and immunological parameters and the antibody response, including the opsonophagocytosis analysis of the sera were evaluated. Moreover the composition of peripheral blood leukocyte populations was studied in infected animals. The results show that the immunization of piglets with rSsnA elicits a significant humoral antibody response. However, the antibody response is not reflected in protection of pigs that are challenged with a virulent strain in our conventional vaccination model. Further studies are necessary to evaluate the use of rSsnA as a vaccine candidate for swine.

Comparison of two biochemical methods for identifying Corynebacterium pseudotuberculosis isolated from sheep and goats

The biochemical pattern of Cowan and Steel (BPCS) was compared with a commercial biochemical strip for the identification of Corynebacterium pseudotuberculosis isolated from small ruminants. On 16S rRNA gene sequencing, 40/78 coryneform isolates from the lymph nodes of sheep and goats with lesions resembling caseous lymphadenitis were identified as C. pseudotuberculosis. The sensitivities of the BPCS and the commercial biochemical strip relative to 16S rRNA sequencing were 80% and 85%, and their specificities were 92.1% and 94.7%, respectively; the level of agreement between the BPCS and the commercial biochemical strip was high (κ=0.82). Likelihood ratios for positive and negative results were 10.0 and 0.22 for the BPCS, and 16.0 and 0.16 for the commercial biochemical strip, respectively. These results indicate that the BPCS and the commercial biochemical strip are both useful for identifying C. pseudotuberculosis in veterinary microbiology laboratories.

Seroprevalence and risk factors of exposure to caprine arthritis-encephalitis virus in southern Spain

The aim of the present transversal descriptive study was to determine the exposure and risk factors associated with caprine arthritis-encephalitis virus (CAEV) in southern Spain. A total of 3,312 serum samples were collected from goat belonging to three different breeds coming from 48 flocks located in different geographic areas from southern Spain. In addition, health and productive parameters were surveyed during the visit to the herds. Serum samples were analysed by INgezim Maedi Screening (Ingenasa®) ELISA kit. The total percentage of herds exposed to CAEV was 87.71% (CI95 78.42–97.00). A total of 733 goats were seropositive with overall seroprevalence of 23.22% (CI95 21.78–24.65). The intraherd seroprevalence was 20.82%±24.07. Multivariate logistic regression showed significant association between CAEV and the next variables: (i) herd size (P<0.0001; OR: 2.07; CI95: 1.73–2.50), (ii) kidding area (P<0.0020; OR: 1.38; CI95: 1.13–1.69), (iii) cleaning and disinfection program (P<0.0067; OR: 1.90; CI95: 1.43–2.53), (iv) natural mating (P<0.0026; OR: 2.22; CI95: 1.73–2.86) and (v) multiparous (P<0.0001; OR: 2.90; CI95: 2.17–3.87). The results indicate a widespread of CAEV in goat herds in southern Spain, with herd size, existence of kidding area, absence of a cleaning and disinfection programme, natural mating and multiparous goats being risk factors for exposure to CAEV.

Antimicrobial activity of selected essential oils against Streptococcus suis isolated from pigs

The inhibitory potential by contact and vapor of basil, cinnamon, clove, peppermint, oregano, rosemary, common thyme, and red thyme essential oils (EOs) against 20 strains of Streptococcus suis was determined by the disk diffusion test. The broth microdilution method was used to determine the minimal inhibitory and minimal bactericidal concentration (MIC and MBC) of the four selected oils. Furthermore, the bactericidal power (ratio MBC/MIC) was calculated. The EOs with the major potential in the disk diffusion method were red thyme, common thyme, oregano, and cinnamon (∅ mean 16.5–34.2 mm), whereas cinnamon did not show vapor activity. In the microdilution test, all the EOs showed notable antimicrobial activity (MIC90 and MBC90 312.5–625 μg·ml−1) and a strong bactericidal power (ratio = 1). This is the first study that selects essential oils against S. suis. New studies about the possible synergic effect of EOs with antibiotics and about toxicity and efficacy in in vivo conditions are recommended.