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    The Prebiotic Effect of an Organic Acid Mixture on Faecalibacterium prausnitzii Metabolism and Its Anti-Pathogenic Role against Vibrio parahaemolyticus in Shrimp
    (MDPI, 2022-12-29) Butucel, Eugenia; Balta, Igori; McCleery, David; Marcu, Adela; Stef, Ducu; Pet, Ioan; Callaway, Todd; Stef, Lavinia; Corcionivoschi, Nicolae
    Increasing the abundance of probiotic bacteria in the gut requires either direct dietary supplementation or the inclusion of feed additives able to support the growth of beneficial commensal bacteria. In crustaceans, the increased presence of probiotic-like bacteria in the gut, including of Faecalibacterium prausnitzii (F. prausnitzii), will guarantee a positive health status and a gut environment that will ensure enhanced performance. The aim of this study was to investigate if a mixture of organic acids, AuraAqua (Aq) can stimulate the growth and the anti-pathogenic efficacy of F. prausnitzii through a combination of in vitro and ex vivo models. The results showed that 0.5% Aq was able to improve the growth rate of F. prausnitzii in vitro and in an ex vivo shrimp gut model. Moreover, we were able to demonstrate that Aq increases butyrate production and cellulose degradation in culture or in the shrimp gut model. The growth-stimulating effect of Aq also led to an improved and anti-pathogenic effect against Vibrio parahaemolyticus in a co-culture experiment with shrimp gut primary epithelial cells (SGP). In conclusion, our work demonstrates that Aq can stimulate the growth of F. prausnitzii, increase the production of short-chain fatty acid (SCFA) butyrate, improve substrate digestion, and prevent V. parahaemolyticus invasion of SGP cells.
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    The Effect Citrox BCL on Legionella pneumophila Mechanisms of Biofilm Formation, Oxidative Stress and Virulence
    (MDPI, 2022-11-04) Butucel, Eugenia; Balta, Igori; McCleery, David; Popescu, Cosmin Alin; Iancu, Tiberiu; Pet, Ioan; Marcu, Adela; Horablaga, Nicolae-Marinel; Stef, Lavinia; Corcionivoschi, Nicolae
    Legionella pneumophila is responsible for causing Legionnaires’ disease and Pontiac fever, also known as legionellosis. The aim of this study was to investigate the mechanistic effect of a mixture of natural antimicrobials (Citrox BCL) in preventing L. pneumophila biofilm formation and reducing its in vitro virulence. The minimum inhibitory concentrations were detected at 0.06%, and the MBC was established at 0.125%. Based on the growth curve profile, the sub-inhibitory concentration of 0.02% was further used to study the mechanistic implications in the absence of a cytotoxic effect on A549 cells. At 24 h post-infection, Citrox BCL reduced (p = 0.005) the intracellular growth of L. pneumophila when the A549 cells or the bacteria were pre-treated with 0.02% Citrox BCL. This result was replicated when Citrox BCL was added during the 24 h infection assay leading to a reduction in intracellular growth (p = 0.003). Herein we show that at the sub-inhibitory concentration of 0.02%, Citrox CBL lowers the ROS levels in infected A549 cells and causes a 45% reduction in L. pneumophila EPS production, a reduction associated with the decline in biofilm formation. Overall, our results corroborate the low c-di-GMP production with the decrease in biofilm formation and low EPS levels. The low EPS levels seemed to be caused by the downregulation of the tatB and tatC gene expressions. Moreover, inhibition of pvcA and pvcB gene expressions, leading to lower siderophore levels, suggests that Citrox BCL reduces the ability of L. pneumophila to sequester iron and reduce biofilm formation through iron starvation.
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    In silico investigation of the genus Campylobacter type VI secretion system reveals genetic diversity in organization and putative effectors.
    (Microbiology Society, 2022-10-31) Robinson, Luca; Liaw, Janie; Omole, Zahra; Corcionivoschi, Nicolae; Hachani, Abderrahman; Gundogdu, Ozan
    Bacterial type VI secretion systems (T6SSs) are contractile nanomachines that deliver proteinic substrates into target prokaryotic or eukaryotic cells and the surrounding milieu. The genus Campylobacter encompasses 39 recognized species and 13 subspecies, with many belonging to a group known as ‘emerging Campylobacter pathogens’. Within Campylobacter, seven species have been identified to harbour a complete T6SS cluster but have yet to be comparatively assessed. In this study, using systematic bioinformatics approaches and the T6SS-positive Campylobacter jejuni 488 strain as a reference, we explored the genus-wide prevalence, similarity and make-up of the T6SS amongst 372 publicly available ‘complete’ Campylobacter genomes. Our analyses predict that approximately one-third of Campylobacter species possess a T6SS. We also putatively report the first identification of a T6SS in four species: Campylobacter cuniculorum, Campylobacter helveticus, Campylobacter armoricus and Campylobacter ornithocola. The Campylobacter T6SSs cluster into three distinct organizations (I–III), of which two break down into further variants. Thirty T6SS-containing genomes were found to harbour more than one vgrG gene, with Campylobacter lari strain NCTC 11845 possessing five. Analysis of the C. jejuni Pathogenicity Island-1 confirmed its conservation amongst T6SS-positive C. jejuni strains, as well as highlighting its diverse genetic composition, including additional putative effector–immunity pairs (e.g. PoNe and DUF1911 domains). Effector–immunity pairs were also observed neighbouring vgrGs in several other Campylobacter species, in addition to putative genes encoding nucleases, lysozymes, ATPases and a ferric ATP-binding cassette uptake system. These observations highlight the diverse genetic make-up of the T6SS within Campylobacter and provide further evidence of its role in pathogenesis.
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    Effect of reduced dietary protein level on finishing pigs’ harmful social behaviour before and after an abrupt dietary change.
    (Elsevier, 2022-10-06) McAuley, Melanie; Buijs, Stephanie; Muns Vila, Ramon; Gordon, Alan Wesley; Palmer, Maeve; Meek, Kayleigh; O'Connell, Niamh
    Tail biting in pigs is a harmful behavioural disorder that negatively affects their welfare. Nutrition has played a crucial role in improving farm efficiency; however, an imbalance in key nutrients is an accepted risk factor for tail biting. The exact contributing factors and the level of risk associated with inadequate nutrition remain unclear. We aimed to assess the effect of dietary protein level on harmful social behaviours in finishing pigs before and after an abrupt dietary change. A total of 80 pigs (Duroc x (Landrace x Large White)) were assigned to this trial over two batches. Pigs were housed in mixed sex groups of 10 and assigned to one of two treatments at 12 weeks of age. These consisted of a control treatment (CON) containing 15.5% Crude Protein (CP) and a low CP treatment (LowCP) containing 13.5% CP. The abrupt dietary change occurred at 16 weeks of age (71.45 kg) in batch 1 and at 15 weeks of age (67.04 kg) in batch 2. This change in diet led to a 1.2% drop in CP in the CON treatment and a 1.4% drop in CP in the LowCP treatment. Behavioural observations were performed before and after the diet change and largely focused on the pigs' social behaviour. Using a "hurdle" model analysis, we investigated whether diet, diet change or sex influenced: 1) the probability that a behaviour was not performed, and 2) the frequency and/or duration when the behaviour was performed. The LowCP treatment led to an increased duration of limb-directed behaviour (P = 0.03) when performed at all, and also an increased duration of ‘tail-in-mouth’ behaviour (P = 0.02) in males but not females. However, LowCP did not affect the level of ear-directed behaviours in this way (P > 0.05). An increase in standing frequency occurred after the abrupt diet change (P = 0.04). Our study indicates that a small reduction in dietary protein and sex affect behaviour in the finishing phase, with some impacts on important harmful social behaviours. Furthermore, an effect of abrupt diet change on behaviour is suggested, which will require further confirmation.
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    The Effect of Transportation on Puppy Welfare from Commercial Breeding Kennels to a Distributor.
    (MDPI, 2022-12-01) Romaniuk, Aynsley C.; Diana, Alessia; Barnard, Shanis; Weller, Jennifer; Espinosa, Uri Baqueiro; Dangoudoubiyam, Sriveny; Shreyer, Traci; Arnott, Gareth; Croney, Candace
    Many puppies from commercial breeding kennels (CBKs) are transported by ground from their kennels of origin to a distributor. This experience may elicit fear and stress during a sensitive developmental period, which may in turn negatively impact the puppies’ short- and long-term welfare. This study aimed to measure short-term effects of transportation on puppy welfare metrics. Eight-week-old puppies (n = 383) from 12 CBKs were tested at their kennels (pre-trans) and ~48 h after arriving at a distributor (post-trans). At each location, puppies underwent an isolation test, a stranger-approach test, and a physical health assessment. Behavioral responses to testing were scored from videos. Fecal glucocorticoid metabolites (FGM), fecal secretory immunoglobulin A (sIgA), and presence of intestinal parasites were also analyzed. Linear mixed-effects models identified decreased exploration (p < 0.001), and increased locomotion (p < 0.001) and escape attempts (p = 0.001) during the post-trans isolation test. Increased affiliative behavior (p < 0.001), FGM (p < 0.001) and sIgA (p = 0.014) were also observed post-trans. Findings support good physical health both pre- and post-trans, while behavioral and physiological changes suggest increased puppy distress post-trans. Higher post-transport affiliative behavior may indicate that puppies sought social support as a coping strategy after experiencing transport-related distress. Future studies should explore the efficacy of transportation-related interventions to mitigate puppy distress.