Biofilms as promoters of bacterial antibiotic resistance and tolerance

Multidrug resistant bacteria are a global threat for human and animal health. However, they are only part of the problem of antibiotic failure. Another bacterial strategy that contributes to their capacity to withstand antimicrobials is the formation of biofilms. Biofilms are associations of microorganisms embedded a self-produced extracellular matrix. They create particular environments that confer bacterial tolerance and resistance to antibiotics by different mechanisms that depend upon factors such as biofilm composition, architecture, the stage of biofilm development, and growth condi-tions. The biofilm structure hinders the penetration of antibiotics and may prevent the accumulation of bactericidal concentrations throughout the entire biofilm. In addition, gradients of dispersion of nutrients and oxygen within the biofilm generate different metabolic states of individual cells and favor the development of antibiotic tolerance and bacterial persistence. Furthermore, antimicrobial resistance may develop within biofilms through a variety of mechanisms. The expression of efflux pumps may be induced in various parts of the biofilm and the mutation frequency is induced, while the presence of extracellular DNA and the close contact between cells favor horizontal gene transfer. A deep understanding of the mechanisms by which biofilms cause tolerance/resistance to antibiotics helps to develop novel strategies to fight these infections

Salmonella infection in nursery piglets and its role in the spread of salmonellosis to further production periods

Few studies have focused on assessing Salmonella infection in the nursery and its role in further pig production periods. Mesenteric lymph nodes, intestinal content, and meat juice from 389 6-week-old male piglets intended for human consumption from five breeding farms and 191 pooled floor fecal samples from gilt development units (GDU) from the same farms were analyzed to estimate and characterize (by pulsed-field gel electrophoresis and antimicrobial resistance analyses) Salmonella infection. The prevalence of infection and shedding among piglets was 36.5% and 37.3%, respectively, shedding being significantly associated with infection (Odds Ratio = 12.7; CI 7.3–22.0). Salmonella Rissen; S. 4, [5], 12:i:-; and S. Derby were the most common serotypes. A low level of Salmonella-specific maternal antibodies at the beginning of the nursery period suggested it was a period of high risk of infection. Resistance to 3rd-and 4th-generation cephalosporins was detected in piglet isolates although the piglets never received antibiotics, indicating they could be vectors of antimicrobial resistance. The same Salmonella clones were detected in piglet and GDU isolates, suggesting that infected piglets play a significant role in the infection of gilts and consequently of finishing pigs in the case of production farms. The control of Salmonella infection in nursery piglets may decrease the risk of abattoir and carcass contamination

Resistance to colistin and production of extended-spectrum ß-lactamases and/or AmpC enzymes in Salmonella isolates collected from healthy pigs in Northwest Spain in two periods: 2008-2009 and 2018

Salmonellosis is a common subclinical infection in pigs and therefore apparently healthy animals may represent a reservoir of antibiotic-resistant Salmonella for humans. This study estimates and characterizes resistance to two classes of antimicrobials considered of the highest priority within the critically important antimicrobials for humans, i.e. colistin (CR) and 3rd generation cephalosporins (3GC), on a collection of Salmonella isolates from pigs from two periods: between 2008 and 09, when colistin was massively used; and in 2018, after three years under a National Plan against Antibiotic Resistance. Prevalence of CR was low (6 out of 625; 0.96%; 95%CI: 0.44–2.1) in 2008–09 and associated mostly to the mcr-1 gene, which was detected in four S. 4,5,12:i:- isolates. Polymorphisms in the pmrAB genes were detected in a S. 9,12:-:- isolate. No CR was detected in 2018 out of 59 isolates tested. Among 270 Salmonella isolates considered for the assessment of resistance to 3GC in the 2008–2009 sampling, only one Salmonella Bredeney (0.37%; 95%CI: 0.07–2.1) showed resistance to 3GC, which was associated with the blaCMY-2 gene (AmpC producer). In 2018, six isolates out of 59 (10.2%; 95%CI: 4.7–20.5) showed resistance to 3GC, but only two different strains were identified (S. 4,12:i:- and S. Rissen), both confirmed as extended-spectrum β-lactamases (ESBL) producers. The blaCTX-M-3 and blaTEM-1b genes in S. 4,12:i:- and the blaTEM-1b gene in S. Rissen seemed to be associated with this resistance. Overall, the prevalence of CR in Salmonella appeared to be very low in 2008–2009 despite the considerable use of colistin in pigs at that time, and seemed to remain so in 2018. Resistance to 3GC was even lower in 2008–2009 but somewhat higher in 2018. Resistance was mostly coded by genes associated with mobile genetic elements. Most serotypes involved in these antimicrobial resistances displayed a multidrug resistance pattern and were considered zoonotic

Evaluation of three commercial enzyme-linked imuunosorbent assays for the detection of antibodies against Salmonella spp. in meat juice from finishing pigs in Spain

The control of animal salmonellosis is considered as a major objective in Europe and indirect ELISAs will be important tools for the implementation of control programs for this infection in pigs. We analyse the results yielded by three commercial ELISAs (Herdcheck Swine Salmonella, SALMOTYPE Pig Screen, and PrioCHECK Salmonella) on meat juice samples from a population of slaughter pigs of Aragon, NW Spain, to assess their efficacy using traditional and latent-class approaches. Overall, the Herdcheck Swine Salmonella detected more Salmonella-infected pigs than the other two tests, but its relative sensitivity was low (65.9%). A similar result was observed when only serotypes detectable by this test were considered (69.1%). When a Bayesian approach was used the Herdcheck Swine Salmonella showed also the highest overall accuracy (sensitivity = 88% and specificity = 74%). Our results suggest that a relatively small proportion of the observed prevalence in herds would be explained by using these ELISAs. Also, this study points out that when different ELISA tests are used within the same herd, results may differ substantially. Thus, caution is advised if it is decided to use these assays for herd health classification in Spanish Salmonella control program

Looking for new approaches for the use of serology in the context of control programmes against pig salmonellosis

Most swine Salmonella national control programmes in Europe have been based on the categorization of herds according to risk levels based on serological results. However, none of the non-Scandinavian countries have reported of any significant success on Salmonella infection reduction in fattening pigs or the number of human cases attributable to pigs or pork. The limited accuracy of the tests used, the small number of animals sampled and the likely lack of herd representativeness of the samples used could be major factors affecting the suitability of these programmes. Focusing on minimizing Salmonella shedding at slaughter appears more important to prevent human infections than focusing on detection of seropositive pigs/herds at this stage. This study assessed whether performing on-farm serology may help to predict shedding at slaughter. Between 2010 and 2016, pigs from six cohorts from a Salmonella-positive herd were bled at 30, 60 and 90 days on fattening and before slaughter, and faecal samples collected at slaughter. Serology on days 60, 90 and before slaughter predicted somewhat shedding at slaughter with no significant differences among them. Pigs with higher OD% values at these point times would have higher risk of shedding when arriving to slaughter. The probability of shedding for a pig sampled on day 90 and showing an OD% value of 10 was 43%, and the risk increased up to 65% if the OD% was 40. Concluding, on-farm serology may help to determine to some extent the risk of Salmonella shedding at slaughter from seropositive fattening units, which would allow for prompt on-farm and slaughter interventions to reduce the likelihood of slaughter contamination with Salmonella.Fil: Mainar-Jaime, R.C.. Universidad de Zaragoza; EspañaFil: Casanova-Higes, A.. Universidad de Zaragoza; EspañaFil: Andrés-Barranco, S.. Universidad de Zaragoza; EspañaFil: Vico, Juan Pablo. Universidad Católica de Córdoba; Argentina. Universidad Católica de Córdoba. Instituto de Investigaciones en Recursos Naturales y Sustentabilidad José Sanchez Labrador S. J. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Recursos Naturales y Sustentabilidad José Sanchez Labrador S. J.; Argentin

Assessment of performance of selected serological tests for diagnosing brucellosis in pigs.

Swine brucellosis due to Brucella suis is considered an emerging zoonotic disease whose control is based on serological testing and the subsequent culling of seropositive animals or the full depopulation of affected flocks. Here we assessed the performance of several serological tests (Rose Bengal Test [RBT], indirect ELISA [i-ELISA], blocking ELISA [b-ELISA], and two competitive ELISAs [c-ELISA]) for diagnosing swine brucellosis caused by B. suis biovar 2. Both frequentistic and Bayesian statistical inference were used. A frequentistic analysis, using sera from known gold standard (GS) populations (i.e., from truly infected or brucellosis free animals), resulted in maximum (100%) diagnostic sensitivity (Se) and specificity (Sp) in the RBT, i-ELISA and b-ELISA tests. However, c-ELISAs resulted in lower diagnostic Se (ranging from 68.5% to 92.6%, according to the different cut-offs selected). A Bayesian analysis of tests yielding the best diagnostic performance with GS sera (RBT, i-ELISA and b-ELISA), but using a large collection of field sera, resulted in similar Se among tests but markedly lower (˜80%) than that resulting from the frequentistic analysis using the GS serum populations. By contrast, the estimated Sp in the Bayesian analysis was only slightly lower than 100%, thus similar to that obtained frequentistically. Our results show that adequate diagnostic tests for brucellosis in swine are available, but also emphasize the need for more extensive validation studies before applying these tests under field conditions