Aims: To test the efficacy of Lactobacillus johnsonii FI9785 in reducing the colonization and shedding of Salmonella enterica serotype Enteritidis, Escherichia coli O78:K80 and Clostridium ...perfringens in poultry.
Methods and Results: Specific pathogen‐free chicks (1 day old) were dosed with a single oral inoculum of 1 × 109 CFU. Lactobacillus johnsonii FI9785 and 24 h later were challenged in separate experiments with S. Enteritidis (S1400, nalr) and E. coli O78:K80 (EC34195, nalr). There were no significant effects against S. Enteritidis whereas colonization of the small intestine by E. coli O78:K80 was reduced significantly. Both S. Enteritidis and E. coli colonized the caeca and colon to levels equivalent to control birds and there was no reduction in shedding as assessed by a semi‐quantitative cloacal swabbing technique. Specific pathogen‐free chicks (20 day old) were dosed with a single oral inoculum of 1 × 109 CFU L. johnsonii FI9785 and 24 h later were challenged with C. perfringens. A single oral dose of L. johnsonii FI9785 was sufficient to suppress all aspects of colonization and persistence of C. perfringens.
Conclusions: Lactobacillus johnsonii FI9785 may be given to poultry for use as a competitive exclusion agent to control C. perfringens.
Significance and Impact of the Study: Lactobacillus johnsonii FI9785 may be a valuable tool to control the endemic disease of necrotic enteritis, thereby reducing economic losses associated with reduced use of antimicrobials in the poultry industry.
► We setup a 3D cells culture able to support HEV viral replication. ► HEV RNA was detected by real time-RT-PCR during all the course of the experiments. ► HEV RNA progeny was detectable by real time ...RT-PCR demonstrating the viral replication. ► HEV like particle were detected by EM.
Hepatitis E is an acute, viral hepatitis epidemic in developing regions, but which is detected with increasing frequency in sporadic form in developed regions. Pigs and possibly some other mammals are considered reservoirs of zoonotic infection with hepatitis E virus (HEV). However, whilst the relative significance of potential transmission routes from pigs to people is still unclear, the consumption of raw or undercooked pig meat has been implicated as a source of HEV infection. The lack of information about HEV zoonotic transmission is due in part to the difficulties of in vitro propagation of HEV. The Rotating Wall Vessel (RVW) has been described as a useful tool for the culture of cell lines in a 3-dimensional (3D) configuration. The aim of this work was to develop a 3D cell culture system for HEV to facilitate studies into the viability of virions contaminating pig tissues. This study, demonstrated that HEV can replicate efficiently in the RWV in human hepatoblastoma PLC/PRF/5 cells for up to 5 months not only by real time RT-PCR but also by detection of complete virions via electron microscopy. Furthermore, the replication of HEV progeny was observed by detecting HEV RNA by RT-PCR. The progeny were able to infect fresh 3D cultures, showing that this method is able to produce infectious hepatitis E virions.
Summary
Escherichia coli comprises a highly diverse group of Gram‐negative bacteria and is a common member of the intestinal microflora of humans and animals. Generally, such colonization is ...asymptomatic; however, some E. coli strains have evolved to become pathogenic and thus cause clinical disease in susceptible hosts. One pathotype, the Shiga toxigenic E. coli (STEC) comprising strains expressing a Shiga‐like toxin is an important foodborne pathogen. A subset of STEC are the enterohaemorrhagic E. coli (EHEC), which can cause serious human disease, including haemolytic uraemic syndrome (HUS). The diagnosis of EHEC infections and the surveillance of STEC in the food chain and the environment require accurate, cost‐effective and timely tests. In this review, we describe and evaluate tests now in routine use, as well as upcoming test technologies for pathogen detection, including loop‐mediated isothermal amplification (LAMP) and whole‐genome sequencing (WGS). We have considered the need for improved diagnostic tools in current strategies for the control and prevention of these pathogens in humans, the food chain and the environment. We conclude that although significant progress has been made, STEC still remains an important zoonotic issue worldwide. Substantial reductions in the public health burden due to this infection will require a multipronged approach, including ongoing surveillance with high‐resolution diagnostic techniques currently being developed and integrated into the routine investigations of public health laboratories. However, additional research requirements may be needed before such high‐resolution diagnostic tools can be used to enable the development of appropriate interventions, such as vaccines and decontamination strategies.
AIMS: The aims of this work were to develop a model of dairy farm waste milk and to investigate methods for the bioremediation of milk containing cefquinome residues. METHODS AND RESULTS: ...Unpasteurized milk and UHT milk that had both been spiked with cefquinome at a concentration of 2 μg ml⁻¹were used as a model for waste milk containing cephalosporin residues. Adjustment of the spiked UHT milk to pH 10 or treatment with conditioned medium from bacterial growth producing cefotaximase, were the most effective methods for decreasing the cefquinome concentrations within 24 h. A large‐scale experiment (10 l of cefquinome‐spiked unpasteurized milk) suggested that fermentation for 22 h at 37°C followed by heating at 60°C for 2 h was sufficient to decrease cefquinome concentrations to below the limit of quantification (<125 μg kg⁻¹) and to kill the majority of the enriched bacterial population. CONCLUSIONS: One or a combination of the bioremediation methods described may have potential as a practical treatment for dairy farm waste milk. SIGNIFICANCE AND IMPACT OF THE STUDY: Treatment of waste milk to decrease cephalosporin residue concentrations and also to kill bacteria prior to feeding to dairy calves could decrease the risk of selection for ESBL bacteria on dairy farms.
A candidate live vaccine for avian pathogenic Escherichia coli (APEC) was constructed from a virulent field APEC O78 strain by mutation of the aroA gene. The mutant was highly similar to the parent ...wild-type strain in respect of colony morphology, motility, growth in suspension, hemagglutination, Congo Red binding, HEp-2 cell adhesion, and the elaboration of surface antigens type 1 fimbriae and flagella, although production of curli fimbriae was reduced marginally. The mutant proved avirulent when inoculated into 1-day-old chicks by spray application and when presented again in the drinking water at 7 days of age. Chickens and turkeys vaccinated with an O78 aroA mutant were protected against a challenge at 6 wk of age by virulent APEC strains. Eficacia de una vacuna viva atenuada de Escherichia coli O78∶K80 en pollos y pavos. Se elaboró una vacuna viva para Escherichia coli patógena aviar (APEC) a partir de una cepa de campo patógena para las aves y virulenta O78 mediante mutación del gene aroA. La mutante fue muy similar a la cepa silvestre original con respecto a la morfología de la colonia, motilidad, crecimiento en suspensión, hemaglutinación, afinidad al rojo Congo, adhesión a células HEp-2 y la elaboración de antígenos de superficie tipo 1 fimbria y flagelo, aunque la producción de fimbrias rizadas se redujo marginalmente. La mutante no mostró virulencia cuando se inoculó en pollitos de un día vía aerosol, ni cuando se administró nuevamente a los 7 días de edad, vía agua de bebida. Los pollos y pavos vacunados con la mutante O78 aroA fueron protegidos contra el desafío a las 6 semanas de edad con una cepa de E. coli patógena aviar virulenta.
The development of novel intervention strategies for the control of zoonoses caused by bacteria such as Salmonella spp. in livestock requires appropriate experimental models to assess their ...suitability. Here, a novel porcine intestinal in vitro organ culture (IVOC) model utilizing cell crown (CC) technology (CCIVOC) (Scaffdex) was developed. The CCIVOC model was employed to investigate the characteristics of association of S. enterica serovar Typhimurium strain SL1344 with porcine intestinal tissue following exposure to a Lactobacillus plantarum strain. The association of bacteria to host cells was examined by light microscopy and electron microscopy (EM) after appropriate treatments and staining, while changes in the proteome of porcine jejunal tissues were investigated using quantitative label-free proteomics. Exposure of porcine intestinal mucosal tissues to L. plantarum JC1 did not reduce the numbers of S. Typhimurium bacteria associating to the tissues but was associated with significant (P < 0.005) reductions in the percentages of areas of intestinal IVOC tissues giving positive staining results for acidic mucins. Conversely, the quantity of neutrally charged mucins present within the goblet cells of the IVOC tissues increased significantly (P < 0.05). In addition, tubulin-α was expressed at high levels following inoculation of jejunal IVOC tissues with L. plantarum. Although L. plantarum JC1 did not reduce the association of S. Typhimurium strain SL1344 to the jejunal IVOC tissues, detection of increased acidic mucin secretion, host cytoskeletal rearrangements, and proteins involved in the porcine immune response demonstrated that this strain of L. plantarum may contribute to protecting the pig from infections by S. Typhimurium or other pathogens.
The potential of a prebiotic oligosaccharide lactulose, a probiotic strain of Lactobacillus plantarum, or their synbiotic combination to control postweaning colibacillosis in pigs was evaluated using ...an enterotoxigenic Escherichia coli (ETEC) K88 oral challenge. Seventy-two weanlings were fed four diets: a control diet (CTR), that diet supplemented with L. plantarum (2 × 10(10) CFU · day(-1)) (LPN), that diet supplemented with 10 g · kg(-1) lactulose (LAC), or a combination of the two treatments (SYN). After 7 days, the pigs were orally challenged. Six pigs per treatment were euthanized on days 6 and 10 postchallenge (PC). Inclusion of lactulose improved the average daily gain (ADG) (P < 0.05) and increased lactobacilli (P < 0.05) and the percentage of butyric acid (P < 0.02) in the colon. An increase in the ileum villous height (P < 0.05) and a reduction of the pig major acute-phase protein (Pig-MAP) in serum (P < 0.01) were observed also. The inclusion of the probiotic increased numbers of L. plantarum bacteria in the ileum and colon (P < 0.05) and in the total lactobacilli in the colon and showed a trend to reduce diarrhea (P = 0.09). The concentrations of ammonia in ileal and colonic digesta were decreased (P < 0.05), and the villous height (P < 0.01) and number of ileal goblet cells (P < 0.05) increased, at day 10 PC. A decrease in plasmatic tumor necrosis factor alpha (TNF-α) (P < 0.01) was also seen. The positive effects of the two additives were combined in the SYN treatment, resulting in a complementary synbiotic with potential to be used to control postweaning colibacillosis.
Spores from a number of different Bacillus species are currently being used as human and animal probiotics, although their mechanisms of action remain poorly understood. Here we describe the ...isolation of 237 presumptive gut-associated Bacillus spp. isolates that were obtained by heat and ethanol treatment of fecal material from organically reared broilers followed by aerobic plating. Thirty-one representative isolates were characterized according to their morphological, physiological, and biochemical properties as well as partial 16S rRNA gene sequences and screening for the presence of plasmid DNA. The Bacillus species identified included B. subtilis, B. pumilus, B. licheniformis, B. clausii, B. megaterium, B. firmus, and species of the B. cereus group, whereas a number of our isolates could not be classified. Intrinsic properties of potential importance for survival in the gut that could be advantageous for spore-forming probiotics were further investigated for seven isolates belonging to five different species. All isolates sporulated efficiently in the laboratory, and the resulting spores were tolerant to simulated gastrointestinal tract conditions. They also exhibited antimicrobial activity against a broad spectrum of bacteria, including food spoilage and pathogenic organisms such as Bacillus spp., Clostridium perfringens, Staphylococcus aureus, and Listeria monocytogenes. Importantly, the isolates were susceptible to most of the antibiotics tested, arguing that they would not act as donors for resistance determinants if introduced in the form of probiotic preparations. Together, our results suggest that some of the sporeformers isolated in this study have the potential to persist in or transiently associate with the complex gut ecosystem.
Cost effective control of avian diseases and food borne pathogens remains a high priority for all sectors of the poultry industry with cleansing and disinfection, vaccination and competitive ...exclusion approaches being used widely. Previous studies showed that
Bacillus subtilis PY79
hr was an effective competitive exclusion agent for use in poultry to control avian pathogenic
Escherichia coli serotype O78:K80. Here we report experiments that were undertaken to test the efficacy of
B. subtilis PY79
hr in the control of
Salmonella enterica serotype Enteritidis and
Clostridium perfringens in young chickens. To do this, 1-day-old and 20-day-old specific pathogen free (SPF) chicks were dosed with a suspension of
B. subtilis spores prior to challenge with
S. Enteritidis (S1400) and
C. perfringens, respectively. For both challenge models, a single oral inoculum of 1×10
9 spores given 24
h prior to challenge was sufficient to suppress colonisation and persistence of both
S. Enteritidis and
C. perfringens. In particular, the faecal shedding of
S. Enteritidis, as measured by a semi-quantitative cloacal swabbing technique, was reduced significantly for the 36 days duration of the experiment.
B. subtilis persisted in the intestine although with decreasing numbers over the same period. These data add further evidence that
B. subtilis spores may be effective agents in the control of avian diseases and food borne pathogens.
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