Phenotypic and genotypic antibiotic resistance patterns in Salmonella Typhimurium and its monophasic variant from pigs in southern Spain.

The high incidence of human salmonellosis and multi-drug resistant (MDR) strains of Salmonella Typhimurium (ST) is of concern to global public and animal health. Our research, by means of the broth microdilution method, evaluated the Minimum Inhibitory Concentration (MIC) distribution of 12 antimicrobials against a collection of 73 ST and mST and S. typhimurium monophasic variant 4,[5],12:i:- (mST) isolates from slaughtered pigs reared in extensive systems in southern Spain, and also 12 resistance-associated genes or antimicrobial resistance (AMR) determinants using qPCR. Our data revealed that 98.6% of strains were MDR, with resistance to cephalothin/tetracycline/sulfamethoxazole-trimethoprim/ampicillin/chloramphenicol being the most common pattern (55.6%). Regarding AMR determinants, the most significantly (p < 0.05) genes detected by qPCR were sul1 and aadA2 (89% of strains positive), aadA1 and dfrA12 (87.7%), and blaTEM and tet(B) (86.3% and 84.9%, respectively). Up to date information on ST antimicrobial resistance patterns is essential for epidemiological surveillance programs to support animal and public health. The high number of MDR isolates and variability regarding resistance determinants revealed in this study highlights the role of animals reared in extensive systems as a source of resistant Salmonella strains.

Serosurvey on schmallenberg virus and selected ovine reproductive pathogens in culled ewes from southern Spain.

After the first case of Schmallenberg virus (SBV) was reported in southern Spain (March 2012), a retrospective serological study was carried out in extensive sheep flocks from nearby areas to assess the history of exposure to SBV and other selected ovine reproductive pathogens (Chlamydophila abortus, Coxiella burnetii, Border Disease virus 'BDV', Toxoplasma gondii and Neospora caninum). Secondly, the presence of antibodies was investigated in meat juice samples against selected pathogens to validate their use in serosurveys in sheep. A total of 209 Merina and cross-bred culled ewes belonging to 12 outdoor flocks managed in extensive breeding systems were sampled. Serum and meat juice samples were collected at the slaughterhouse and analysed using commercial ELISA kits. Chlamydophila abortus (62.68%, CI95 56.13-69.23) and Toxoplasma gondii (57.42%, CI95 50.72-64.12) were the most prevalent pathogens. The seroprevalence of BDV (16.27%, CI95 11.27-21.27) and Coxiella burnetii (13.88%, CI95 9.2-18.56) was moderate, and only 4 of 209 animals (1.91%, CI95 1.82-2.96) presented specific antibodies against Neospora caninum or SBV. All the examined ovine flocks were seropositive to three or more pathogens. The highest percentage of seropositive animals was detected for T. gondii-C. abortus coseropositive (25.36%) culled ewes. The concordance between serum and meat juice samples was moderate for T. gondii (κ = 0.419) and BDV (κ = 0.568), and fair for C. abortus (κ = 0.311). Our results show evidence of circulation of SBV from summer 2011 in southern Spain. Furthermore, C. abortus and T. gondii were the most prevalent pathogens associated with sheep in outdoor rearing systems. Finally, these preliminary results point to meat juice samples as a potential biological sample for serosurveys studies on sheep.

A serological survey of Brucella spp., Salmonella spp., Toxoplasma gondii and Trichinella spp. in Iberian fattening pigs reared in free-range systems.

Zoonotic agents such as Brucella spp., Salmonella spp., Toxoplasma gondii and Trichinella spp., all considered high-risk zoonotic pathogens by the European Food Safety Agency (EFSA), may cause no symptoms of infection in free-range pigs yet still have a significant public health impact. A serological survey was therefore performed to determine the history of occurrence of these pathogens in such pigs in southern Spain. A total of 709 serum samples were collected at abattoir from pigs from 79 farms and analysed for specific antibodies against the above pathogens using commercially available ELISA kits. Encysted Trichinella spp. larvae were also sought following the artificial digestion method of diaphragm pillar muscle. The results showed Salmonella spp. to be widely distributed among the sampled herds [73.42%, 95% confidence interval (CI95 ) 65.6-81.78] and Toxoplasma gondii to be present in over half (58.23%, CI95 47.33-69.07). The seroprevalence of Brucella spp. was very low (3.8%, CI95 0.18-7.42), and antibodies against Trichinella spp. were not detected. No encysted Trichinella spp. larvae were microscopically detected.

Salmonella Indiana as a cause of abortion in ewes: Genetic diversity and resistance patterns.

Salmonella enterica subspecies enterica Indiana, a food-borne serovar uncommon in most countries, was responsible for an outbreak of abortion in a flock of Lacaune dairy ewes in southern Spain. Drinking water and feedstuff samples were analysed in an attempt to determine the source of the infection. Pigeons (Columba livia) and turtledoves (Streptopelia turtur) in close contact with the ewes were captured and examined for the bacterium. Seventeen S. Indiana strains were isolated from the ewes and wild birds and the genetic similarity among them analysed by Pulsed Field Gel Electrophoresis (PFGE) after the digestion of their genomic DNA with the restriction enzyme XbaI. The results suggest the wild birds might be responsible for the outbreak in the ewes. The strains recovered were fully susceptible to 15 out of the 16 antimicrobial agents tested: ampicillin, amoxycillin clavulanate, cephalothin, ceftriaxone, gentamicin, neomycin, streptomycin, tetracycline, ciprofloxacin, enrofloxacin, sulphonamides, trimethoprim-sulphamethoxazole, apramycin, colistin and chloramphenicol. Differences in the resistance pattern to nalidixic acid were observed; 11 strains (64.7%) were nalidixic acid resistant (R-Nx) and 6 (35.3%) sensitive (S-Nx). Among the R-Nx strains, a substitution of Gly to Cys at position 81 (Gly81àCys) of the gyrA gene in 10 strains isolated from wild birds and ovine foetuses, and of Asp to Tyr at position 87 (Asp87àTyr) in one strain isolated from ewe faeces, were revealed by sequencing the gene. To control the outbreak, enrofloxacin treatment was administered for 5 days. The same therapy was used to prevent infection during following gestation cycles, administering the antimicrobial agent at presentation and over 4 weeks before birth. Anti-bird meshes and closed drinking and feeding troughs were also installed to prevent further contact of the ewes with wild birds.

Pneumonic pasteurellosis associated with Pasteurella haemolytica in chipmunks (Tamias sibiricus).

A laboratory study was performed in order to identify the possible cause of death in chipmunks (Tamias sibiricus) imported from China with respiratory disease. Severe congestion, alveolar oedema and fibrinous pleuritis were observed. Biochemical analyses identified the causative organism as Pasteurella haemolytica. An in vitro susceptibility test using various antimicrobial agents revealed sensitivity to beta-lactams (ampicillin and amoxicillin) and streptomycin.

Avian influenza in wild waterfowl and shorebirds in the Donana National Park: Serological survey using the enzyme-linked immunosorbent assay.

The indirect ELISA was used to detect antibodies to influenzavirus A in the sera of wildfowl from the Donana National Park. Of the 712 birds examined, 44 (6.2%) were seropositive. Positive birds belonged to 10 of the 13 species studied. Infection rates varied widely: spoonbill (Platalea leucorodia, 32.2%), mallard (Anas platyrhynchos, 9.9%), gadwall (Anas strepera, 8.6%), red-crested pochard (Netta rufina, 8.1%), pochard (Aythya ferina, 6.4%), shoveler (Anas clypeata, 5%), great crested grebe (Podiceps cristatus, 4.3%), avocet (Recurvirostra avosetta, 3.1%), grey heron (Ardea cinerea, 3.1%) and coot (Fulica atra, 0.8%). Although infection rates were not high, the wide range of avian species susceptible to influenzavirus A suggests circulation of the virus amongst wildfowl at Donana.

Serological survey of infections in waterfowl in the Guadalquivir marshes (Spain).

A serological survey was performed in the Guadalquivir Marshes (Southern Spain) in order to determine the presence and diffusion of six infective agents in wild waterfowl. The analysis covered 712 waterfowl from 13 species belonging to five taxa (Podicipediformes, Ciconiiformes, Anseriformes, Gruiformes, and Charadriiformes). A range of immunological techniques led to detection of antibodies against the six infective agents studied: Chlamydia psittaci (13.3%), Mycoplasma anatis (3.5%), Pasteurella anatipestifer (2.9%), duck plague virus (1.6%), Aspergillus fumigatus (1.1%), and Newcastle disease virus (NDV) (0.2%). The survey results indicated that these infective agents exist and circulate among wild waterfowl in the Guadalquivir Marshes.