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et al. ; Guano samples from 412 Brazilian bats were screened with real-time PCR for the virulence genes (eae, est, elt, stx1, stx2, ehxA, invA, bfpA, aggR) representing five intestinal pathotypes of Escherichia coli. From 82 pooled samples, 22% contained Escherichia coli DNA, and eae, est, bfpA, aggR were detected. ; Research was supported by European Union FP7 ANTIGONE (Anticipating the Global Onset of Novel Epidemics) project 278976 ; Peer Reviewed

SIMPLE SUMMARY: Spanish production of compound feed is among the most important in the Member States of the European Union for all livestock species. However, due to the environmental impact of this large-scale production system, it is important to focus on sustainability, promoting initiatives such as the use of by-products from the food industry applied to animal feed. In this study, laying hens received two types of dietary supplement: biscuit meal, which is a co-product of the human food industry commonly used in the manufacture of compound feed, obtained from the recycling of wasted or expired food products; and fermented defatted "alperujo", a by-product of modified olive oil, which contain numerous substances with beneficial properties for intestinal health. Hens co-administered with these supplements showed increased intestinal villi development, resulting in improved health. In conclusion, these by-products can contribute to the prevention of intestinal diseases, as well as to the reduction of environmental pollution. ABSTRACT: In this study, the effects of co-administration with biscuit flour and fermented defatted "alperujo" (FDA) on gut health were evaluated in a batch of laying hens (Hy-Line 2015) on a commercial farm. Animals were divided into two groups: control group and treatment group; and histological and morphometric analyses of all sections of the intestine (duodenum, jejunum, ileum, cecum and rectum) were performed at 10, 18, 25, 50 and 75 weeks of age. During the whole productive period, a decrease in the mortality rate (p = 0.01) was observed in treated hens, as well as an increase in the number of eggs produced (p < 0.001), their size (p < 0.025), and weight (p < 0.024). In the early and late stages of production (10, 18 and 50 weeks), a significant increase (p < 0.001) in the height and depth of the intestinal villi was observed in the treatment group. Villi height was also significantly higher (p < 0.001) in the treatment group up to week 50 in the cecum, and at weeks 18 ...

South American camelids are susceptible to tuberculosis, caused mainly by Mycobacterium bovis and M. microti. Despite the tuberculin skin test being the official test for tuberculosis, it has a very low sensitivity in these species (14-20%). Serological tests present the advantages of being rapid, easy to perform and facilitate analysis of large numbers of samples in a short period of time. Novel antigen discovery and evaluation would provide enhanced detection of specific antibodies against members of M. tuberculosis complex. Here, we describe the development and evaluation of an ELISA-type immunoassays to use in the diagnosis of tuberculosis in llamas and alpacas based on P22, a multiprotein complex obtained by affinity chromatography from bovine Purified Protein Derivative (bPPD), that showed high sensitivity and specificity in mice, cattle and goats. This work was performed in two stages. First, a preliminary panel of samples collected from tuberculosis-free (n = 396) and M. bovis-infected herds (n = 56) was assayed, obtaining high specificity (100%) and sensitivity ranging from 63 to 96%. Subsequently, the use of the serological assay was tested using samples from two herds suffering from clinical M. bovis (n = 88) and M. microti (n = 25) infection to evaluate the ability of the ELISA to detect infected animals. 11 out of 88 alpacas were positive to the ELISA in a M. bovis outbreak and 7 out of 25 in a M. microti outbreak. The P22 ELISA potentially provides a sensitive and specific platform for improved tuberculosis surveillance in camelids. ; JI-L was supported by an FPU contract-fellowship (Formación de Profesorado Universitario) from the Ministerio de Educación, Cultura y Deporte of the Spanish Government (FPU2013/6000). AR is the recipient of an Industrial Doctorate contract (DI-15-08110) funded by the Spanish Ministry of Economy, Industry and Competitiveness (MINECO) and the European Social Fund. This work was funded by the University of Surrey Innovation Voucher Scheme 2017-18 ; Sí

SIMPLE SUMMARY: Salmonella spp. is a bacterium that places human health at risk by consuming eggs and poultry. In the European Union, the use of antimicrobials to treat salmonellosis in aviculture is no longer permitted due to the resistance to treatment of some bacteria, such as Salmonella spp. For this reason, compounds derived from natural food sources are being increasingly tested to assess their efficacy against Salmonella spp. In this study, chickens were given dietary supplements in the form of fermented defatted 'alperujo', a modified olive oil by-product, after which they were infected with Salmonella Typhimurium. The chickens given the supplement showed a healthy gut and a reduction in the amount of Salmonella spp. in the cecum. In conclusion, this olive oil by-product may contribute to preventing and controlling salmonellosis in farms, as well as reducing environmental contamination. ABSTRACT: Salmonella spp. contaminates egg and poultry meat leading to foodborne infections in humans. The emergence of antimicrobial-resistant strains has limited the use of antimicrobials. We aimed to determine the effects of the food supplement, fermented defatted 'alperujo' (FDA), a modified olive oil by-product, on Salmonella Typhimurium colonisation in broilers. One hundred and twenty 1-day-old broilers were divided into four experimental groups—two control groups and two treated groups, and challenged with S. Typhimurium at day 7 or 21. On days 7, 14, 21, 28, 35, and 42 of life, duodenum and cecum tissue samples were collected for histopathological and histomorphometric studies. Additionally, cecum content was collected for Salmonella spp. detection by culture and qPCR, and for metagenomic analysis. Our results showed a significant reduction of Salmonella spp. in the cecum of 42-day-old broilers, suggesting that fermented defatted 'alperujo' limits Salmonella Typhimurium colonization in that cecum and may contribute to diminishing the risk of carcass contamination at the time of slaughter. The improvement of the ...

European badgers (Meles meles ) have been identified as wildlife reservoirs for Mycobacterium bovis in the UK and Ireland, and may also have a role in the epidemiology of animal tuberculosis in other European regions. Thus, detection of M. bovis‐infected badgers may be required for the purposes of surveillance and monitoring of disease levels in infected populations. Current serological assays to detect M. bovis infection in live badgers, while rapid and inexpensive, show limited diagnostic sensitivity. Here we describe and evaluate new ELISA platforms for the recognition of the P22 multiprotein complex derived from the purified protein derivative (PPD ) of M. bovis . The recognition of IgG against P22 multiprotein complex derived from PPD ‐B was tested by ELISA in the serum of badgers from the UK , Ireland and Spain. TB infection in the badgers was indicated by the presence of M. bovis in tissues by culture and histology at post‐mortem examination and TB ‐free status was established by repeated negativity in the interferon γ release assay (IGRA ). In experimentally infected badgers, humoral antibody responses against P22 developed within 45 days post‐infection. The ELISA tests showed estimated sensitivity levels of 74–82% in experimentally and naturally infected badgers with specificities ranging from 75% to 100% depending on the badger population tested. The P22 multi‐antigen based ELISA s provide a sensitive and specific test platform for improved tuberculosis surveillance in badgers. ; This work was supported by the Instituto Nacional de Investigacion y Tecnologia Agraria y Alimentaria of Spain (INIA; RTA2015-00043-C02-02) and the TAVS-CM Programme of the Comunidad de Madrid (S2013/ABI-2747), cofinanced by the FEDER fund 'A way to build Europe'. This work was partially supported by a FEDER co-funded grant from INIA (RTA2014-00002-C02-01). Jose Antonio Infantes Lorenzo was supported by an FPU contract-fellowship (Formacion de Profesorado Universitario) from the Ministerio de Educacion, Cultura y Deporte of the Spanish Government (FPU2013/6000). ; Peer reviewed

Animal tuberculosis (TB) remains a major problem in some countries despite the existence of control programmes focused mainly on cattle. In this species, aerogenous transmission is accepted as the most frequent infection route, affecting mainly the respiratory system. Under the hypothesis that the oral route could be playing a more relevant role in transmission, diagnosis and disease persistence than previously thought, this study was performed to assess the course of TB infection in cattle and its effects on diagnosis depending on the route of entry of Mycobacterium bovis. Two groups of five calves each were either endotracheally (EC) or orally (OC) challenged. Necropsies were carried out 12 weeks after challenge except for three OC calves slaughtered 8 weeks later. All animals reacted to the tuberculin skin test and the entire EC group was positive to the interferon-gamma release assay (IGRA) 2 weeks after challenge and thereafter. The first positive IGRA results for OC calves (3/5) were recorded 4 weeks after challenge. Group comparison revealed significant differences in lesion and positive culture location and scoring. TB-compatible gross lesions and positive cultures were more frequently found in the thorax (p < 0.001) and lung (p < 0.05) of EC animals, whereas OC animals presented lesions (p = 0.23) and positive cultures (p < 0.05) mainly located in the abdomen. These results indicate that the infection route seems to be a determining factor for both the distribution and the time needed for the development of visible lesions. Our study suggests that confirmation of TB infection in some skin reactor animals can be problematic if current post-mortem examination and diagnostics are not improved. ; This study was supported with funds from the Spanish Ministry of Economy and Competitiveness (Research Project AGL2014-56305-C3-3-R) and the Department of Economic Development and Competitiveness of the Basque Government. MS holds a fellowship from the Department of Education of the Basque Government (PRE_2017_2_0043). ; Peer Reviewed

We found SARS-CoV-2 RNA in 6 of 71 ferrets (8.4%) and isolated the virus from one rectal swab. Natural SARS-CoV-2 infection does occur in kept ferrets, at least under circumstances of high viral circulation in the human population. However, small ferret collections are probably unable to maintain prolonged virus circulation. ; This study received funding from ISCIII, Spanish Government, grant number: COV20/01385 and EFC was supported by a grant from Universidad de Castilla-La Mancha, Spain. ; No

Eight seafood protein hydrolysates (SPHs) obtained from squid, shrimp and fish gelatin were incorporated as substitutes of peptones in culture media in order to evaluate its effect on survival and metabolic activity (lactic acid, acetic acid and bacteriocins production) of Enterococcus faecalis DM19. The substitution of commercial peptones in culture media by either a shrimp hydrolysate prepared with Protamex, or by squid protein hydrolysates prepared with Esperase or Alkaline protease, stimulated E. faecalis DM19 growth up to 16%. The incorporation of SPHs, mainly from shrimp, in the culture media significantly increased production of lactic and acetic acids in more than 60%. Furthermore, the media containing SPHs stimulated antimicrobial activity by E. faecalis DM19. The inhibitory activity was observed against both Gram-positive and Gram-negative microorganisms, but it was remarkably observed against Listeria monocytogenes. SPHs incorporated in culture media render properties of bio-technological interest, which, together with their low price, make them suitable for industrial use. ; This research was financed by the Spanish Ministry of Economy and Competitiveness (projects AGL2011-27607, AGL2014-52825-R). Author M. Djellouli is funded by The National Centre of Biotechnology Research (CHER-Stage 06-2013) (CNRBt) of Algeria and ENP (Exceptional National Program) Scholarship provided by the Ministry of Higher Education and Scientific Research of Algeria (099/PNE/ENS./ESPAGNE/2015-2016). Author M. Arancibia is funded by a SENESCYT Scholarship (20100338) provided by the Ecuadorian government. ; Peer Reviewed

The ante-mortem diagnosis of tuberculosis (TB) in ruminants is based mainly on the intradermal tuberculin test and the IFN-γ assay. Antibody (Ab)-based tests have emerged as potential tools for the detection of TB infected animals using serum, plasma, or even milk samples. Oral fluids have also been evaluated as alternative samples with which to detect specific Abs against Mycobacterium bovis in pigs or wild boars, but not in ruminants. The objective of this study was, therefore, to evaluate the performance of an in house-ELISA for TB diagnosis (P22 ELISA) in goats as an experimental model for the diagnosis of TB using oral fluid samples. Oral fluid samples from 64 goats from a TB-infected herd (n = 197) and all the animals from a TB-free herd (n = 113) were analyzed using the P22 ELISA. The estimated sensitivity (Se) and specificity (Sp) were 34.4% (95% CI: 22.4-45.6) and 100% (95% CI: 97.4-100), respectively. The optimal cut-off point was set at 100% according to the ROC analysis. Those animals with a higher level of Abs in their oral fluid attained a higher lesion score (p = 0.018). In fact, when taking into account only the setting of the animals with severe lesions (n = 16), the ELISA showed a Se of 75% (95% CI: 53.7-96.2). Results of the present study suggest that the P22 ELISA is highly specific but has a limited value detecting infected animals in oral fluid samples. Nevertheless, its performance is significantly higher in the presence of severe lesions. ; This study was funded by the Herramientas para alcanzar la erradicación de la tuberculosis caprina (GoaTBfree) project (PID2019-105155RB-C31) and the Spanish Government's Ministerio de Agricultura, Pesca y Alimentación. JO was supported by an FPU (Formación de Profesorado Universitario) contract-fellowship provided by the Spanish Ministerio de Ciencia, Innovación y Universidades (FPU18/05197). ; Sí

This is an open access article distributed under the terms of the Creative Commons Attribution License.-- et al. ; Mycobacteria of the Mycobacterium tuberculosis complex (MTBC) greatly affect humans and animals worldwide. The life cycle of mycobacteria is complex and the mechanisms resulting in pathogen infection and survival in host cells are not fully understood. Recently, comparative genomics analyses have provided new insights into the evolution and adaptation of the MTBC to survive inside the host. However, most of this information has been obtained using M. tuberculosis but not other members of the MTBC such as M. bovis and M. caprae. In this study, the genome of three M. bovis (MB1, MB3, MB4) and one M. caprae (MB2) field isolates with different lesion score, prevalence and host distribution phenotypes were sequenced. Genome sequence information was used for whole-genome and protein-targeted comparative genomics analysis with the aim of finding correlates with phenotypic variation with potential implications for tuberculosis (TB) disease risk assessment and control. At the whole-genome level the results of the first comparative genomics study of field isolates of M. bovis including M. caprae showed that as previously reported for M. tuberculosis, sequential chromosomal nucleotide substitutions were the main driver of the M. bovis genome evolution. The phylogenetic analysis provided a strong support for the M. bovis/M. caprae clade, but supported M. caprae as a separate species. The comparison of the MB1 and MB4 isolates revealed differences in genome sequence, including gene families that are important for bacterial infection and transmission, thus highlighting differences with functional implications between isolates otherwise classified with the same spoligotype. Strategic protein-targeted analysis using the ESX or type VII secretion system, proteins linking stress response with lipid metabolism, host T cell epitopes of mycobacteria, antigens and peptidoglycan assembly protein identified new genetic markers and candidate vaccine antigens that warrant further study to develop tools to evaluate risks for TB disease caused by M. bovis/M.caprae and for TB control in humans and animals. ; This research was supported by grants AGL2014-56305 and IPT-2011-0735-010000 from Ministerio de Economía y Competitividad, Spain, and the European Union FP7 ANTIGONE grant 278976 and Horizon 2020 COMPARE Grant 377/14. ; Peer Reviewed

[Background]: Bovine purified protein derivative (bPPD) and avian purified protein derivative (aPPD) are widely used for bovine tuberculosis diagnosis. However, little is known about their qualitative and quantitative characteristics, which makes their standardisation difficult. In addition, bPPD can give false-positive tuberculosis results because of sequence homology between Mycobacterium bovis (M. bovis) and M. avium proteins. Thus, the objective of this study was to carry out a proteomic characterisation of bPPD, aPPD and an immunopurified subcomplex from bPPD called P22 in order to identify proteins contributing to cross-reactivity among these three products in tuberculosis diagnosis. [Methods]: Trypsin digests of bPPD, aPPD and P22 were analysed by nanoscale liquid chromatography-electrospray ionization tandem mass spectrometry. Mice were immunised with bPPD or aPPD, and their serum was tested by indirect ELISA for reactivity against these preparations as well as against P22. [Results]: A total of 456 proteins were identified in bPPD, 1019 in aPPD and 118 in P22; 146 of these proteins were shared by bPPD and aPPD, and 43 were present in all three preparations. Candidate proteins that may cause cross-reactivity between bPPD and aPPD were identified based on protein abundance and antigenic propensity. Serum reactivity experiments indicated that P22 may provide greater specificity than bPPD with similar sensitivity for ELISA-type detection of antibodies against M. tuberculosis complex. [Conclusion]: The subpreparation from bPPD called P22 may be an alternative to bPPD for serodiagnosis of bovine tuberculosis, since it shares fewer proteins with aPPD than bPPD does, reducing risk of cross-reactivity with anti-M. avium antibodies. ; This work was supported by the Ministerio de Economía, Industria y Competitividad of Spain (RTC-2016-4746-2), the Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria of Spain (RTA2015-00043-C02-02) and the TAVS-CM Programme of the Comunidad de Madrid (S2013/ABI-2747), cofinanced by the FEDER fund "A way to build Europe". Jose Antonio Infantes-Lorenzo was supported by an FPU contract-fellowship (Formación de Profesorado Universitario) from the Ministerio de Educación, Cultura y Deporte of the Spanish Government (FPU2013/6000). Alvaro Roy is the recipient of an Industrial Doctorate contract (DI-15-08110) funded by the Spanish Ministerio de Economía, Industria y Competitividad and the European Social Fund. ; Peer Reviewed

Background: Bovine purified protein derivative (bPPD) and avian purified protein derivative (aPPD) are widely used for bovine tuberculosis diagnosis. However, little is known about their qualitative and quantitative characteristics, which makes their standardisation difficult. In addition, bPPD can give false-positive tuberculosis results because of sequence homology between Mycobacterium bovis (M. bovis) and M. avium proteins. Thus, the objective of this study was to carry out a proteomic characterisation of bPPD, aPPD and an immunopurified subcomplex from bPPD called P22 in order to identify proteins contributing to cross-reactivity among these three products in tuberculosis diagnosis. Methods: Trypsin digests of bPPD, aPPD and P22 were analysed by nanoscale liquid chromatography-electrospray ionization tandem mass spectrometry. Mice were immunised with bPPD or aPPD, and their serum was tested by indirect ELISA for reactivity against these preparations as well as against P22. Results: A total of 456 proteins were identified in bPPD, 1019 in aPPD and 118 in P22; 146 of these proteins were shared by bPPD and aPPD, and 43 were present in all three preparations. Candidate proteins that may cause cross-reactivity between bPPD and aPPD were identified based on protein abundance and antigenic propensity. Serum reactivity experiments indicated that P22 may provide greater specificity than bPPD with similar sensitivity for ELISA-type detection of antibodies against M. tuberculosis complex. Conclusion: The subpreparation from bPPD called P22 may be an alternative to bPPD for serodiagnosis of bovine tuberculosis, since it shares fewer proteins with aPPD than bPPD does, reducing risk of cross-reactivity with anti-M. avium antibodies. ; This work was supported by the Ministerio de Economía, Industria y Competitividad of Spain (RTC-2016-4746-2), the Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria of Spain (RTA2015-00043-C02-02) and the TAVS-CM Programme of the Comunidad de Madrid (S2013/ABI-2747), cofinanced by the FEDER fund "A way to build Europe". Jose Antonio Infantes-Lorenzo was supported by an FPU contract-fellowship (Formación de Profesorado Universitario) from the Ministerio de Educación, Cultura y Deporte of the Spanish Government (FPU2013/6000). Alvaro Roy is the recipient of an Industrial Doctorate contract (DI-15-08110) funded by the Spanish Ministerio de Economía, Industria y Competitividad and the European Social Fund. ; Sí