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Within the global endeavours to reduce antibiotic resistances and protect human and animal health, novel antimicrobials are urgently required (WHO, 2022a). In the WHO report on clinical development against priority pathogens 2022, nine out of 34 non-traditional antimicrobials were bacteriophages or bacteriophage-based products (WHO, 2022b). Bacteriophages (phages) are the natural enemies of bacteria, they can infect their host bacteria, replicate inside the bacterial cell and finally kill bacteria by lysis. Their therapeutic potential for reducing bacterial pathogens in birds has been shown in several studies (KITTLER et al., 2017). Due to their mode of action, phages meet all innovation criteria of the WHO, belonging to a new chemical class, having new molecular targets and showing no cross resistance to other antibiotic classes. They have been used against Campylobacter in several in vivo trials and two field trials in commercial broiler houses (KITTLER et al., 2013; CHINIVASAGAM et al., 2020). While phage resistance in Campylobacter was associated with fitness costs in some experiments, the occurrence of new bacterial strains that are not sensitive to the bacteriophages used, was suggested to reduce phage effectiveness and reproducibility under field conditions. Thus, phages were combined with other measures in a multiple-hurdle approach in a seeder bird model (PEH et al., 2024) and under field conditions (BOGUN et al., 2024). The application of a Fletchervirus phage NCTC 12673 and Firehammervirus phage vB_CcM-LmqsCPL1/1 in a field trial, resulted in reductions of up to 1.1 log10 CFU/ml in fecal samples 1 day after dosing. Combined application of the phages with curcumin via feed and an organic acid blend via drinking water did not result in higher reductions than phage application alone. Based on these results, no synergism of the combined application was observed. However, after combined application of the phages with a competitive exclusion culture in a seeder bird model, increased reduction compared to single used measures has been observed. [ABSTRACT FROM AUTHOR]

Aim: The family Arcobacteraceae formerly genus Arcobacter has recently been reclassified into six genera. Among nine species of the genus Aliarcobacter, Aliarcobacter faecis and Aliarcobacter lanthieri have been identified as emerging pathogens potentially cause health risks to humans and animals. This study was designed to develop/optimize, validate and apply Arcobacteraceae family‐ and two species‐specific (A. faecis and A. lanthieri) loop‐mediated isothermal amplification (LAMP) assays to rapidly detect and quantify total number of cells in various environmental niches. Methods and Results: Three sets of LAMP primers were designed from conserved and variable regions of 16S rRNA (family‐specific) and gyrB (species‐specific) genes. Optimized Arcobacteraceae family‐specific LAMP assay correctly amplified and detected 24 species, whereas species‐specific LAMP assays detected A. faecis and A. lanthieri reference strains as well as 91 pure and mixed culture isolates recovered from aquatic and faecal sources. The specificity of LAMP amplification of A. faecis and A. lanthieri was further confirmed by restriction fragment length polymorphism analysis. Assay sensitivities were tested using variable DNA concentrations extracted from simulated target species cells in an autoclaved agricultural water sample by achieving a minimum detection limit of 10 cells mL−1 (10 fg). Direct DNA‐based quantitative detection, from agricultural surface water, identified A. faecis (17%) and A. lanthieri (1%) at a low frequency compared to family‐level (93%) with the concentration ranging from 2·1 × 101 to 2·2 × 105 cells 100 mL−1. Conclusions: Overall, these three DNA‐based rapid and cost‐effective novel LAMP assays are sensitive and can be completed in less than 40 min. They have potential for on‐site quantitative detection of species of family Arcobacteraceae, A. faecis and A. lanthieri in food, environmental and clinical matrices. Significance and Impact of the Study: The newly developed LAMP assays are specific, sensitive, accurate with higher reproducibility that have potential to facilitate in a less equipped lab setting and can help in early quantitative detection and rate of prevalence in environmental niches. The assays can be adopted in the diagnostic labs and epidemiological studies. [ABSTRACT FROM AUTHOR]

The loop‐mediated isothermal amplification (LAMP) technique was used to investigate six salmonella‐specific sequences for their suitability to serve as targets for the pathogen identification. Sequences selected for designing LAMP primers were genes invA, bcfD, phoP, siiA, gene62181533 and a region within the ttrRSBCA locus. Primers including single nucleotide polymorphisms were configured as degenerate primers. Specificity of the designed primer sets was determined by means of 46 salmonella and 32 other food‐ and waterborne bacterial reference species and strains. Primers targeting the ttrRSBCA locus showed 100 % inclusivity of target and exclusivity of other test species and strains. Other primer sets revealed deficiencies, especially regarding Salmonella enterica subsp. II–IV and Salmonella bongori. Additionally, primers targeting the siiA gene failed to detect S. enterica subsp. enterica serotypes Newport and Stanley, whereas bcfD primers did not amplify DNA of S. enterica subsp. enterica serotype Schleissheim. TtrRSBCA primers, providing short detection times and constant melting temperatures of amplification products, achieved best overall performance. [ABSTRACT FROM AUTHOR]

The proapoptotic B-cell lymphoma (Bcl)-2 protein Bcl-xS encloses the Bcl-2 homology (BH) domains BH3 and BH4 and triggers apoptosis via the multidomain protein Bak, however, the mechanism remained elusive. For investigating Bcl-xS efficacy and pathways, an adenoviral vector was constructed with its cDNA under tetracycline-off control. Bcl-xS overexpression resulted in efficient apoptosis induction and caspase activation in melanoma cells. Indicative of mitochondrial apoptosis pathways, Bcl-xS translocated to the mitochondria, disrupted the mitochondrial membrane potential and induced release of cytochrome c, apoptosis-inducing factor and second mitochondria-derived activator of caspases. In melanoma cells, Bcl-xS resulted in significant Bak activation, and Bak knockdown as well as Bcl-xL overexpression abrogated Bcl-xS-induced apoptosis, whereas Mcl-1 (myeloid cell leukemia-1) knockdown resulted in a sensitization. With regard to the particular role of voltage-dependent anion channel 2 (VDAC2) for inhibition of Bak, we identified here a notable interaction between Bcl-xS and VDAC2 in melanoma cells, which was proven in reciprocal coimmunoprecipitation analyses. On the other hand, Bcl-xS showed no direct interaction with Bak, and its binding to VDAC2 appeared as also independent of Bak expression. Suggesting a new proapoptotic mechanism, Bcl-xS overexpression resulted in disruption of the VDAC2-Bak interaction leading to release of Bak. Further supporting this pathway, overexpression of VDAC2 strongly decreased apoptosis by Bcl-xS. New proapoptotic pathways are of principle interest for overcoming apoptosis deficiency of melanoma cells. [ABSTRACT FROM AUTHOR]

Large numbers of infections and effective treatment or control depend on the ability to identify the causative agents of animal infections. Here we describe a study of an isolate DG22/1209/12 obtained from vaginal pus exudate collected from a Siamese cat. The isolate was believed to be a member of Trueperella abortisuis, which is related to problems with reproduction. The most important was to identify this isolate by applying different techniques of diagnosis. It included phenotypic testing, Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS), and molecular techniques such as amplification and sequencing of the 16S rRNA gene, intergenic spacer region (ISR), and the gap and tuf genes. Phenotypic tests confirmed the identification of DG22/1209/12 as T. abortisuis. Finally, the identification was confirmed with MALDI-TOF MS analysis which demonstrated log (score) value of 2.09 indicating a secure genus identification, probable species identification on T. abortisuis. Molecular analysis revealed a high sequence identity of 99.9 %, 99.5 %, 99.4 %, and 99.5 % for the 16S rRNA gene, ISR, gap, and tuf genes, respectively, for both the isolate DG22/1209/12 and the reference isolate T. abortisuis DSM 19515 T . The study highlights the effectiveness of combining phenotypic and genomic methods to accurately identify bacterial pathogens in veterinary medicine, leading towards focused intervention strategies., Competing Interests: Declaration of competing interest There is no conflict of interest among the contributors of this work., (Copyright © 2024. Published by Elsevier B.V.)

Proteins from insect production represent an interesting (environmentally friendly) option or supplement to commercial livestock farming. At present, however, the larval stages of T. molitor (mealworm) and A. diaperinus (buffalo worm) have been authorized as food for human consumption EU-wide, as have the nymph and adult stages of Locusta ( L. ) migratoria ( Locusta migratoria , Linnaeus, 1758) and Acheta ( A. ) domesticus (house cricket, Acheta domesticus , Linnaeus, 1758). However, there is the problem that insects that are recognizable as a whole tend to be avoided by consumers, especially in the European region, as they are reminiscent of living things and can cause aversion and disgust in consumers. Against this background, in the present study, five batches of two types of cooked sausages were produced: on the one hand, with turkey, and on the other hand, with pork lean meat as a base. In different formulations, 10% and 20% of the meat contents (turkey or pork) in these meat products were replaced by deep-frozen, pulverized T. molitor and A. diaperinus larvae. The effects of the addition of these insects in the products on the microbiological and physicochemical parameters of these cooked sausages, compared to a product without insect content, directly after heating, were investigated. After production, a storage trial was also carried out to determine whether possible insect ingredients could influence the growth of inoculated bacterial species ( Bacillus ( B. ) cereus , Escherichia ( E. ) coli , Listeria ( L. ) monocytogenes , and Campylobacter ( C. ) jejuni ) and how the addition of insect larvae affectsthe sensory and physicochemical properties during storage. The study showed that the products with insects had reduced lightness (turkey p C = 0.025), increased yellowness (pork p S = 0.0009, p C < 0.0001 and turkey p C = 0.0027) and a reduced red color (pork p S < 0.0001, p C = 0.0001) after heating when compared to the cooked sausages without insects. However, no significant differences between the various cooked sausages with or without insects in terms of cooking loss, firmness, and protein, ash, and fat or water contents were found. The microbiological tests showed, on the one hand, that the prior microbial reduction (e.g., in the form of blanching) of the insect larvae was essential in order to guarantee the flawless microbiological quality of the cooked sausages and, on the other hand, that the addition of insects to the cooked sausages did not significantly affect the growth of the inoculated bacterial species and that no sensory differences could be detected during storage. Despite the significant color effects on the product, A. diaperinus and T. molitor larvae would be suitable as protein or meat alternatives in cooked sausages, but they would have to undergo pre-treatment, primarily with regard to microbiological safety. The extent to which a complete replacement of meat is possible has to be investigated in further studies., Competing Interests: The authors declare no conflicts of interest.

Streptococcus (S.) equi subspecies (subsp.) zooepidemicus is an important facultative pathogen in horses and can cause severe infections in other species including humans. Facing the post-antibiotic era, novel antimicrobials are needed for fighting bacterial infections. Bacteriophages (phages) are the natural predators of bacteria and discussed as a promising antimicrobial treatment option. The objective of this study was to isolate and characterize S. equi subsp. zooepidemicus- specific phages for the first time and to evaluate their efficacy in vitro and ex vivo . In total, 13 phages with lytic activity were isolated and host ranges were determined. Two phages with broad host ranges and high efficiency of plating (vB&#95;SeqZP&#95;LmqsRe26-2 (lytic activity: 30/37 bacterial isolates) and vB&#95;SeqZP&#95;LmqsRe26-3 (lytic activity: 29/37 bacterial isolates)) and one phage with relatively low efficiency of plating (vB&#95;SeqZP&#95;LmqsRe26-1) were selected for further characterization, including electron microscopy and whole genome sequencing. In in vitro planktonic killing assays at two tested multiplicities of infection (MOI 1 and MOI 10), significant bacterial growth reduction was observed when the phages vB&#95;SeqZP&#95;LmqsRe26-2 and vB&#95;SeqZP&#95;LmqsRe26-3 were added. These phages were subsequently co-incubated with clinical S. equi subsp. zooepidemicus isolates in an equine endometrial explant model but did not achieve bacterial growth reduction at MOI 1 and MOI 10. However, helium ion microscopy revealed presence of particles adherent to the bacteria on the explant after incubation (25 h), suggesting possible phage-bacteria interactions. In conclusion, phages against S. equi subsp. zooepidemicus were successfully isolated and characterized. Promising results were observed in in vitro but no significant reduction was detected in ex vivo experiments, requiring additional investigations. However, after further adaptations (e.g., optimization of MOIs and phage administration or use of phage-antibiotic combination), phages could be a potential antimicrobial tool for future therapeutic use in S. equi subsp. zooepidemicus infections, although the available results do not currently support the therapeutic usage., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Köhne, Hüsch, Tönissen, Schmidt, Müsken, Böttcher, Hirnet, Plötz, Kittler and Sieme.)

The present study investigated the microbiological safety of the increasingly popular plant-based milk alternatives. No (10/27) or only very low microbial counts (17/27) were detected in the tested products. These were mainly identified as spore formers via MALDI-ToF-MS. Three products contained Bacillus cereus group isolates, which were able to form considerable amounts of enterotoxins and exhibited cytotoxicity towards CaCo-2 cells. Preliminary tests showed good growth of B. cereus, Listeria monocytogenes, and Salmonella enterica in all tested products (maximum bacterial counts: 5 × 10 12 cfu/mL). These experiments also revealed strain-, time-, and temperature-, but especially product-specific enterotoxin production of B. cereus. In propagation and persistence tests according to DIN EN ISO 20976-1:2019-09, rapid bacterial proliferation was also detected in all products. B. cereus generally showed lower bacterial counts (10 6 -10 7 cfu/mL) compared to L. monocytogenes and S. enterica (10 8 -10 9 cfu/mL), but was detectable in a larger number of products over the test period of 6 weeks. pH values decreased (20/27) over time and visual and/or olfactory alterations (24/27) were observed. The present study provides information on the occurrence, growth and persistence of pathogenic bacteria in plant-based drinking milk alternatives. It also points out that the accompanying changes in pH, odor, and appearance are not necessarily recognizable to the consumer. PRACTICAL APPLICATION: The present study contributes to the understanding of the microbial risk related to plant-based drinking milk alternatives. It is crucial that the manufacturer ensures that particularly spore formers have been effectively eliminated from the products. Among them, especially toxin-producing bacteria can pose a risk to the consumer, as these products promote proliferation and persistence of the bacteria., (© 2024 The Author(s). Journal of Food Science published by Wiley Periodicals LLC on behalf of Institute of Food Technologists.)

Previous case reports indicate Arcanobacterium 's opportunistic pathogenic potential. However, the true diversity of the genus remains understudied. Here, we present the complete genome of Arcanobacterium wilhelmae isolated from a diseased rhinoceros, suspected to play a role in its condition. These genomic data may enable future advancements in understanding Arcanobacterium pathogenicity., Competing Interests: The authors declare no conflict of interest.

Autoantibodies against contactin-associated protein 2 (Caspr2) not only induce limbic autoimmune encephalitis but are also associated with pain conditions. Here, we analyzed clinical data on pain in a large cohort of patients included into the German Network for Research in Autoimmune Encephalitis. Out of 102 patients in our cohort, pain was a frequent symptom (36% of all patients), often severe (63.6% of the patients with pain) and/or even the major symptom (55.6% of the patients with pain). Pain phenotypes differed between patients. Cluster analysis revealed two major phenotypes including mostly distal-symmetric burning pain and widespread pain with myalgia and cramps. Almost all patients had IgG4 autoantibodies and some additional IgG1, 2, and/or 3 autoantibodies, but IgG subclasses, titers, and presence or absence of intrathecal synthesis were not associated with the occurrence of pain. However, certain pre-existing risk factors for chronic pain like diabetes mellitus, peripheral neuropathy, or preexisting chronic back pain tended to occur more frequently in patients with anti-Caspr2 autoantibodies and pain. Our data show that pain is a relevant symptom in patients with anti-Caspr2 autoantibodies and support the idea of decreased algesic thresholds leading to pain. Testing for anti-Caspr2 autoantibodies needs to be considered in patients with various pain phenotypes., (© 2024. The Author(s).)

In this study, changes in hepatitis E virus (HEV) contamination in the production of liver sausage from naturally contaminated pork liver were investigated. Furthermore, the potential effectiveness of individual production parameters in reducing viral loads was measured. When processing moderately contaminated liver (initial Cq -value 29), HEV RNA persisted in the finished sausages, even after heating for 90 min at 75 °C. A matrix-specific standard curve was created using a spiking experiment to accurately quantify HEV RNA in a particularly challenging matrix like liver sausage. Variations in product-specific production parameters, including mincing and heating times, showed some reduction in contamination levels, but even prolonged heating did not render all finished products HEV negative. The persistence of HEV contamination underscores the importance of ongoing monitoring in the pig population and raw materials to enhance food safety measures and reduce the likelihood of transmission through pork consumption. The detection of HEV RNA within all processing stages of pork liver in the production of liver sausage suggests that further research into the risk of infection posed by this detection and vigilance in managing HEV risks in the food chain, particularly in pork products, are required to protect public health.

Campylobacter mitigation along the food production chain is considered effective for minimizing the public health burden of human campylobacteriosis. This study is the first combining different measures in a multiple-hurdle approach, using drinking water additives and feed additives in single and combined application schemes in commercial broiler plants. Broiler chickens in the study groups were naturally contaminated with Campylobacter. Application of an organic acid blend via drinking water, consisting of sodium propionate, potassium sorbate, and sodium diacetate, resulted in significant reductions of up to 4.9 log 10 CFU/mL in fecal samples and in cecal samples at slaughter. The application of a phage mixture, consisting of Fletchervirus phage NCTC 12673 and Firehammervirus phage vB&#95;CcM-LmqsCPL1/1, resulted in reductions of up to 1.1 log 10 CFU/mL in fecal samples 1 day after dosing. The sole administration of curcumin via feed resulted in small and inconsistent reductions. In the group receiving a combination of all tested measures, reductions of up to 1.1 log 10 CFU/mL were observed. Based on the results of our field trials, it was shown that both the sole application and the combined application of mitigation measures in primary production can reduce the Campylobacter load in broiler chickens, while no synergism could be observed., (© 2024. The Author(s).)

As an international and zoonotic cause of hepatitis, hepatitis E virus (HEV) poses a significant risk to public health. However, the frequency of occurrence and the degree of contamination of food of animal origin require further research. The aim of this study was to develop and validate a highly sensitive quantitative RT-qPCR assay for the detection and quantification of HEV contamination in porcine liver and food. The focus was on genotype 3, which is most common as a food contaminant in developed countries and Europe. The selected assay has its target sequence in the open reading frame 1 (ORF1) of the HEV genome and showed good results in inclusivity testing, especially for HEV genotype 3. The developed assay seems to show high efficiency and a low intercept when compared to other assays, while having a comparable limit of detection (LOD). In addition, a standard curve was generated using artificially spiked liver to provide more accurate quantitative results for contamination assessment and tracking in this matrix. Application of the assay to test 67 pig livers from different origins resulted in a positivity rate of 7.5%, which is consistent with the results of numerous other prevalence studies. Quantitative detection of the viral genome in the food chain, particularly in pig livers, is essential for understanding the presence and evolution of HEV contamination and thus ensures consumer safety.

Many species of the genus Arcanobacterium are known as opportunistic pathogens and have been isolated in association with infectious diseases in humans and animals. Here, we present the complete genome sequence of another opportunistic pathogenic representative, namely Arcanobacterium canis , isolated from the otitis externa of an English bulldog., Competing Interests: The authors declare no conflict of interest.

Meat can be contaminated with (pathogenic) microorganisms during slaughter, dissection and packaging. Therefore, preservation technologies are frequently used to reduce the risk of (fatal) human infections due to the consumption of meat. In this study, we first investigated, if the application of ethyl-Nα-dodecanyl-L-arginate hydrochloride (LAE) and the starter culture bacteria Staphylococcus carnosus and Lactobacillus sakei , either single or in combination, influences the bacteria number on pork, chicken meat and beef, inoculated with Brochothrix (Br.) thermosphacta (all meat species) or Salmonella ( S .) Typhimurium (pork), Campylobacter ( C .) jejuni (chicken) and Listeria ( L .) monocytogenes (beef), before packaging under modified atmosphere and on days 7 and 14 of storage. To evaluate effects of the treatment on the appearance during storage, additionally, the physicochemical parameters color and myoglobin redox form percentages were analyzed. LAE regularly resulted in a significant reduction of the number of all bacteria species on day 1 of storage, whereas up to day 14 of storage, the preservation effect did not persist in nearly all samples, except in the beef with Br. thermosphacta . However, with the starter culture bacteria on day 1, only L. monocytogenes on beef was significantly reduced. Interestingly, on day 7 of storage, this reducing effect was also found with S . Typhimurium on pork. Br. thermosphacta , which was principally not influenced by the starter culture bacteria. The combinatory treatment mainly resulted in no additional effects, except for the S. Typhimurium and Br. thermosphacta results on pork on day 7 and the Br. thermosphacta results on beef on day 14. The physicochemical parameters were not influenced by the single and combinatory treatment. The results indicate that LAE was mainly responsible for the antimicrobial effects and that a combination with starter culture bacteria should be individually evaluated for the meat species., Competing Interests: The authors declare no conflicts of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

The iron-binding glycoprotein lactoferrin is well known for its wide range of antibacterial effects. However, the aim of this study was to show that its antibacterial activity is not generally applicable to a bacterial species as a whole. In disk diffusion assays performed with 112 isolates from 13 bacterial species (including the foodborne pathogens Bacillus cereus and Staphylococcus aureus), a lactoferrin-based food supplement showed no inhibition of growth on 24%, moderate inhibition on 31%, and strong inhibition on 45% of all tested isolates. Minimal inhibitory concentrations against B. cereus and Bacillus thuringiensis strain-specifically ranged from 0.31 mg/mL to no impairment at all. Further 11 commercially available lactoferrin-based food supplements and purified bovine lactoferrin showed strain- as well as product-specific growth inhibition. In comparison to bovine lactoferrin, human lactoferrin showed no inhibitory effects. In summary, purified lactoferrin and lactoferrin-based food supplements inhibit bacterial growth in a dose-, strain-, and product-dependent manner. Thus, a general antimicrobial effect of lactoferrin against a specific bacterial species cannot be assumed., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Aims: To reduce Campylobacter along the food chain, we investigated the mitigation potential of four antimicrobial compounds against Campylobacter using a new evaluation scheme., Methods and Results: Using the checkerboard method, the minimum inhibitory concentration (MIC) values of two organic acids (peroxyacetic acid and lactic acid) and two plant extracts (carvacrol and resveratrol) against a C. jejuni and a C. coli field isolate were determined as well as the fractional inhibitory concentration (FIC) indices of combined treatment. The lowest MIC values were found for peroxyacetic acid (0.03 mg mL-1) and carvacrol (0.06 mg mL-1). Based on subsequent sensory studies, peroxyacetic acid and carvacrol were selected for challenge tests to quantitatively determine the reducing potential against Campylobacter on chicken meat and chicken skin. Applying peroxyacetic acid significantly reduced Campylobacter counts on chicken skin with maximum reductions of 3.3 log-units (P < .0001), while the combination of peroxyacetic acid and carvacrol resulted in significant reductions of only 0.4 log-units on chicken breast fillet 24 hours after treatment but not thereafter (P = .0192)., Conclusions: Peroxyacetic acid is suitable as a postharvest intervention measure to reduce Campylobacter concentration on chicken skin without reducing consumer acceptance., (© The Author(s) 2023. Published by Oxford University Press on behalf of Applied Microbiology International.)

Trueperella pecoris was described as a new species of the genus Trueperella in 2021 and might be pathogenic to various animal species. However, the lack of a suitable diagnostic test system stands in the way of epidemiological surveys to clarify possible causalities. In this study, a Loop-mediated Isothermal Amplification (LAMP) assay was developed and validated that was highly specific for T. pecoris. The assay provided an analytical sensitivity of 0.5 pg/25 µL and showed 100% inclusivity and exclusivity for 11 target and 33 non-target strains, respectively. Three different DNA extraction methods were evaluated to select the most LAMP-compatible method for cell disruption in pure and complex samples. Using an on-site applicable single-buffer DNA extraction with additional heating, the cell-based detection limit was 2.3 CFU/reaction. Finally, the LAMP assay was validated by means of artificially contaminated porcine lung tissue samples in which minimal microbial loads between 6.54 and 8.37 × 10 3  CFU per swab sample were detectable. The LAMP assay established in this study represents a suitable diagnostic procedure for identifying T. pecoris in clinical specimens and will help to collect epidemiological data on the pathogenicity of this species., (© 2023. Springer Nature Limited.)

Background: Effective strategies are urgently needed to control Campylobacteriosis, one of the most important foodborne gastrointestinal diseases worldwide. Administering bacteriophages (phages) is under evaluation as a possible intervention strategy in primary poultry production to reduce the public health risk of human infection. A major challenge is the translation of results from small-scale animal studies to large broiler flocks. In this study, the in vitro lytic activity of 18 Campylobacter-specific group II phages and 19 group III phages were examined singly, and in different combinations from the same group and from both groups using a planktonic killing assay. Based on these results, a combination of phage NCTC 12,673 (group III) and vB&#95;CcM-LmqsCPL1/1 (group II) was selected for in vivo application in a seeder bird model to study its effectiveness under conditions as close as possible to field conditions. One hundred eighty Ross 308 broiler chickens were divided into a control and a treatment group. Ten days post hatch, seeder birds were orally inoculated with the C. jejuni target strain. Phages were administered via drinking water at a total concentration of 10 7 PFU/mL four, three, and two days before necropsy., Results: Combining group II and group III phages resulted in significantly higher in vitro growth inhibition against the C. jejuni target strain BfR-CA-14,430 than single application or combinations of phages from the same group. The results of the animal trial showed that the application of the two phages significantly reduced Campylobacter counts in cloacal swabs. At necropsy, Campylobacter counts in colonic content of the treatment group were significantly reduced by 2 log 10 units compared to the control group., Conclusions: We demonstrated that combining phages of groups II and III results in significantly increased lytic activities. The in vitro results were successfully translated into practical application in a study design close to field conditions, providing new data to apply phages in conventional broiler flocks in the future. Phage application reduced the fecal Campylobacter excretion and Campylobacter concentrations in the colon of broilers., (© 2023. BioMed Central Ltd., part of Springer Nature.)

The use of proteins from insects, plants, microalgae, fungi or bacteria as an alternative to proteins of animal origin such as meat, fish, eggs or milk can meet the worldwide protein demand in the future. As the consumption of whole insects might be problematic or unacceptable for many consumers, especially in European countries, the use of homogenized insects or protein extracts from insects for the production of products might be a possibility to overcome general acceptability problems. However, the quality criteria of these products have to be comparable with consumers' expectations with regard to known products. Therefore, in the present study, we produced a meat product, replaced 10% and 20% of the pork with homogenized larvae of Tenebrio molitor and Hermetia illucens , and determined different physicochemical and sensory parameters at production and during modified atmosphere storage for 21 days. Additionally, the alteration of different bacteria species during this storage was analyzed in challenge tests. After production, the addition of insects resulted in higher cooking losses and pH values in the products with 20% insects, higher pH and yellowness, lower lightness, protein and hardness results in the Hermetia products, as well as higher yellowness and lower protein and hardness values in the cooked meat products with Tenebrio molitor . During modified atmosphere storage, the color differences principally remained, whereas the concentrations of inoculated Bacillus cereus , Listeria monocytogenes and Escherichia coli were not influenced by the addition of insects to the cooked meat products. The sensory results of the insect products, especially at higher concentrations and with Hermetia illucens , worsened during modified atmosphere storage. The addition of homogenized insect larvae, especially at higher concentrations and particularly of Hermetia illucens , influences different physicochemical and sensory parameters of the cooked meat products.

Foodborne diseases are mainly caused by the contamination of meat or meat products with pathogenic microorganisms. In this study, we first investigated the in vitro application of TRIS-buffered plasma-activated water (Tb-PAW) on Campylobacter (C.) jejuni and Escherichia (E.) coli , with a reduction of approx. 4.20 ± 0.68 and 5.12 ± 0.46 log 10 CFU/mL. Furthermore, chicken and duck thighs (inoculated with C. jejuni or E. coli ) and breasts (with natural microflora) with skin were sprayed with Tb-PAW. Samples were packed under a modified atmosphere and stored at 4 °C for 0, 7, and 14 days. The Tb-PAW could reduce C. jejuni on days 7 and 14 (chicken) and E. coli on day 14 (duck) significantly. In chicken, there were no significant differences in sensory, pH-value, color, and antioxidant activity, but %OxyMb levels decreased, whereas %MetMb and %DeoMb increased. In duck, we observed slight differences in pH-value, color, and myoglobin redox forms for the Tb-PAW, which were not perceived by the sensory test persons. With only slight differences in product quality, its application as a spray treatment may be a useful method to reduce C. jejuni and E. coli on chicken and duck carcasses.

The genus Arcanobacterium is constantly growing as novel species are identified. In particular, harbor seals have proven to be a common reservoir for bacteria of this genus. Here, we announce the complete genome sequence of another Arcanobacterium species-namely, Arcanobacterium pinnipediorum strain DSM 28752, isolated from a harbor seal.

Tuna is one of the most widely consumed fish on the European market, being available in various consumable options. Among them, Thunnus albacares, also called yellowfin tuna, is a delicacy and is consumed by millions of people around the world. Due to its comparatively high cost and demand, it is more vulnerable to fraud, where low-cost tuna or other fish varieties might be replaced for economic gain. In this study, a loop-mediated isothermal amplification (LAMP) assay was developed and validated for targeting the mitochondrial cytochrome b gene for fast and direct detection of Thunnus albacares, which is a valuable tuna species. The analytical specificity was confirmed using 18 target samples (Thunnus albacares) and 18 samples of non-target fish species. The analytical sensitivity of the LAMP assay was 540 fg DNA per reaction. In addition, a simple and direct swab method without time-consuming nucleic acid extraction procedures and the necessity for cost-intensive laboratory equipment was performed that allowed LAMP detection of Thunnus albacares samples within 13 minutes. Due to its high specificity and sensitivity, the LAMP assay can be used as a rapid and on-site screening method for identifying Thunnus albacares, potentially providing a valuable monitoring tool for food authenticity control by the authorities., Competing Interests: The authors have declared that no competing interests exist.

Campylobacteriosis is still the most commonly reported zoonosis in the European Union causing gastrointestinal disease in humans. One of the most common sources for these food-borne infections is broiler meat. Interactions between Campylobacter ( C .) jejuni and the intestinal microbiota might influence Campylobacter colonization in chickens. The aim of the present study was to gain further knowledge about exclusive interactions of the host microbiota with C. jejuni in Campylobacter -specific phage-free chickens under standardized conditions and special biosafety precautions.Therefore, 12 artificially infected ( C. jejuni inoculum with a challenge dose of 7.64 log 10 c.f.u.) and 12 control chickens of the breed Ross 308 were kept under special biosafety measures in an animal facility. At day 42 of life, microbiota studies were performed on samples of caecal digesta and mucus. No Campylobacter -specific phages were detected by real-time PCR analysis of caecal digesta of control or artificially infected chickens. Amplification of the 16S rRNA gene was performed within the hypervariable region V4 and subsequently sequenced with Illumina MiSeq platform. R (version 4.0.2) was used to compare the microbiota between C. jejuni -negative and C. jejuni -positive chickens. The factor chickens' infection status contributed significantly to the differences in microbial composition of mucosal samples, explaining 10.6 % of the microbiota variation ( P =0.007) and in digesta samples, explaining 9.69 % of the microbiota variation ( P =0.015). The strongest difference between C. jejuni -non-infected and C. jejuni -infected birds was observed for the family Peptococcaceae whose presence in C. jejuni -infected birds could not be demonstrated. Further, several genera of the family Ruminococcaceae appeared to be depressed in its abundance due to Campylobacter infection. A negative correlation was found between Christensenellaceae R-7 group and Campylobacter in C. jejuni -colonised chickens, both genera potentially competing for substrate. This makes Christensenellaceae R-7 group highly interesting for further studies that aim to find control options for Campylobacter infections and assess the relevance of this finding for chicken health and Campylobacter colonization.

A polyphasic taxonomic study was performed on an unidentified previously described Arcanobacterium -like Gram-positive strain 2701 T isolated from an anal swab of a dead male harbour seal. Comparative 16S rRNA sequencing showed that the bacterium belonged to the genus Arcanobacterium in the family Arcanobacteriaceae . The genome sequence of the strain was obtained by Borowiak et al . [1]. The genome had a G+C content of 49 mol% and a total length of 1.94 Mb. The presence of the major menaquinone MK-9(H 4 ) supported the affiliation of the isolate with the genus Arcanobacterium . The polar lipid profile consisted of diphosphatidylglycerol and an unidentified phospholipid as major components and two unidentified lipids, a further unidentified phospholipid, two unidentified phosphoglycolipids as well as phosphatidylglycerol. The major fatty acids were C 16 : 0 , C 18 : 1 and C 18 : 0 . Biochemical and phylogenetic analyses clearly distinguished the isolate from other members of the genus Arcanobacterium and closely related other species. Based on these results, it is proposed that the unknown Arcanobacterium sp. strain 2701 T should be classified as representing a novel species with the name Arcanobacterium buesumense sp. nov. The type strain is 2701 T (=DSM 112952 T =LMG 32446 T ).

Listeria monocytogenes represents a high risk in food and can trigger potentially fatal listeriosis. The objective of this study was to detect L. monocytogenes in food using the LAMP method in a fast, specific, sensitive manner and thus to preventively test food for the presence of the target species. The reaction was performed and established using the portable real-time fluorometer Genie ® II (OptiGene Ltd., Horsham, United Kingdom). In this new assay, six LAMP primers targeted the mpl -gene sequence of L. monocytogenes . A total of 148 different isolates, including 105 L. monocytogenes and 43 non- L. monocytogenes strains, were tested. Analytical sensitivity was determined based on different DNA- and cell concentrations. The detection limit with a detection rate of 100% was 5 pg of DNA or 275 colony-forming units (CFU) per reaction. Artificially contaminated minced beef and grated mozzarella were also tested. The assay was 100% successful to detect an initial bacterial contamination of 0.4-4 CFU g -1 food after 24 h enrichment in half-Fraser broth. Finally, natively contaminated samples were tested in comparison to the microbiological reference method and real-time polymerase chain reaction. Native sample testing revealed 100% consistent findings between LAMP and the standard culture method after first enrichment for 24 h. In addition, a rapid colony-confirmation method was established that enabled reliable identification of L. monocytogenes isolates on different selective culture media using a simplified DNA extraction by boiling. This study showed that the developed assay was able to determine whether a food is safe with respect to the food-safety criteria of 100 CFU per gram, according to standards of the European Union, for L. monocytogenes and provided faster results than the cultural reference method.

The closely related members of the Bacillus cereus -group can mainly only be differentiated by whole genome sequencing. Among them, there are potentially toxin-producing bacteria. When consumed with food, these can cause vomiting or diarrhea and abdominal cramps. To date, although no EU-wide threshold exists, a bacterial count of 10 5 CFU/g can be regarded as critical. Specific and rapid detection of the bacteria is difficult due to their close relationship, and no loop-mediated isothermal amplification (LAMP) assay has been developed so far to detect potentially toxin-producing members of the B. cereus -group. Aim of this study was to develop a LAMP method to detect critical cell counts specifically and rapidly of potentially non-haemolytic enterotoxin (NHE)-producing cells of this group. A two-step LAMP assay was developed. First, the target sequence groEL was used to determine the representatives of the B. cereus -group. Second, since bacteria in which nheB is present are basically capable of producing enterotoxins, this gene was chosen for detection. The specificity of the developed assay was 100% for B. cereus -group isolates and 93.7% for the detection of nheB . The analytical sensitivity was 0.1 pg DNA/μl. Using simplified DNA extraction by boiling, cell-based sensitivity was determined. Targeting groEL and nheB , 11.35-27.05 CFU/reaction and 11.35-270.5 CFU/reaction were detectable, respectively. Artificially contaminated samples were investigated to prove the application in foods. Direct detection of the critical value of B. cereus -group cells was possible in 83.3% of samples and detecting the toxin-gene 50% thereof. After a 6-h incubation period, the detection rate increased to 100 and 91.7%, respectively. Additionally, 100 natively contaminated food samples were tested, also quantitatively and culturally. Samples with relevant contamination levels were reliably detected using groEL -LAMP. After a 6-h incubation period, isolates bearing the toxin gene nheB could also be reliably detected. In addition, colony material was boiled and used as a LAMP template for simple detection. Specificity for the B. cereus -group was 100 and 93.22% detecting nheB. The study demonstrated that screening of food samples with the groEL / nheB -LAMP assay can be performed within 1 day, making it possible to detect critical levels of potentially NHE-toxin-producing cells of the B. cereus -group., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Busch, Schotte, Jeßberger, Frentzel, Plötz and Abdulmawjood.)

Statements that naturally cured meat products may contain lower residual nitrite levels compared to classical variants led to a closer examination of emulsion-type sausage products in this study, where the input of nitrate from plant extracts (red beet and Swiss chard) was adjusted to typical input levels of nitrite from nitrite curing salt (0.5% NaNO 2 ). The investigations showed that an incubation period of 150 min at 38 °C was necessary to complete the microbial reduction process of nitrate to nitrite and that residual nitrite contents of naturally cured sausages were comparable to the conventionally cured variant, regardless of the nitrate source. During the incubation period, the starter cultures were the dominant microorganisms and showed competitive properties against the natural accompanying flora. In terms of colour development, the variants with Swiss chard juice extract as well as synthetic nitrate showed similar colour formation to conventionally produced emulsion-type sausages. In contrast, colour-providing components of the red beet extract considerably masked the typical appearance., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The national deployment of polyvalent community health workers (CHWs) is a constitutive part of the strategy initiated by the Ministry of Health to accelerate efforts towards universal health coverage in Haiti. Its implementation requires the planning of future recruitment and deployment activities for which mathematical modelling tools can provide useful support by exploring optimised placement scenarios based on access to care and population distribution. We combined existing gridded estimates of population and travel times with optimisation methods to derive theoretical CHW geographical placement scenarios including constraints on walking time and the number of people served per CHW. Four national-scale scenarios that align with total numbers of existing CHWs and that ensure that the walking time for each CHW does not exceed a predefined threshold are compared. The first scenario accounts for population distribution in rural and urban areas only, while the other three also incorporate in different ways the proximity of existing health centres. Comparing these scenarios to the current distribution, insufficient number of CHWs is systematically identified in several departments and gaps in access to health care are identified within all departments. These results highlight current suboptimal distribution of CHWs and emphasize the need to consider an optimal (re-)allocation., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2022 Champagne et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The first description of Trueperella (T.) abortisuis was presented in Japan in 2009 by Azuma and colleagues. In the current study, eight T. abortisuis strains were identified by a newly developed loop-mediated isothermal amplification (LAMP) assay based on the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) encoding gene gap. Two T. abortisuis strains were obtained from prepuce of a seven-month-old boar and pooled foetal stomach contents in the United Kingdom, while the other six T. abortisuis strains were recovered from aborted foetal material of six pigs from a single farm in Germany. The developed LAMP assay showed an analytical sensitivity of 22 pg μL -1 T. abortisuis DNA. T. abortisuis DSM 19515 T and field strain T. abortisuis P504054/19/1 were directly detectable in artificially contaminated vaginal swabs up to concentrations of 980 CFU and 770 CFU per swab, respectively. There was no cross reactivity with control strains representing six species of genus Trueperella and six species of the closely related genus Arcanobacterium and Schaalia (Actinomyces) hyovaginalis. Further field research is required to determine the usefulness of the designed LAMP assay for identifying T. abortisuis isolated from pigs of various origins and from test samples directly obtained at farm level., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Trueperella (T.) bernardiae is a well-known bacterial pathogen in infections of humans, rarely in animals. In the present study, five T. bernardiae isolates, isolated from five Peking ducks of four different farms, were identified by phenotypic properties, by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis, and genotypically by sequencing the 16S ribosomal RNA (rRNA) gene, the superoxide dismutase A encoding gene sodA, and the glyceraldehyde-3-phosphate dehydrogenase encoding gene gap. In addition, the T. bernardiae isolates could be identified with a newly developed loop-mediated isothermal amplification (LAMP) assay based on the gyrase encoding housekeeping gene gyrA. All these tests clearly identified the T. bernardiae isolates to the species level. However, the detection of the specific gene gyrA with the newly designed LAMP assay appeared with a high sensitivity and specificity, and could help to identify this bacterial species in human and animal infections in future. The importance of the T. bernardiae isolates for the clinical condition of the ducks and for the problems at farm level remains unclear., (© 2021. The Author(s).)

Campylobacteriosis is a worldwide-occurring disease and has been the most commonly reported zoonotic gastrointestinal infection in the European Union in recent years. The development of successful phage-based intervention strategies will require a better understanding of phage-bacteria interactions to facilitate advances in phage cocktail design. Therefore, this study aimed to investigate the effects of newly isolated group II and group III phages and their combinations on current Campylobacter field strains. A continuous workflow for host range and efficiency of plating (EOP) value determination was combined with a qPCR-based phage group identification and a liquid-based planktonic killing assay (PKA). An advanced analysis scheme allowed us to evaluate phage cocktails by their efficacy in inhibiting bacterial population growth and the resulting phage concentrations. The results of this study indicate that data obtained from PKAs are more accurate than host range data based on plaque formation (EOP). Planktonic killing assays with Campylobacter appear to be a useful tool for a straightforward cocktail design. Results show that a group II phage vB&#95;CcM-LmqsCP218-2c2 and group III phage vB&#95;CjM-LmqsCP1-1 mixture would be most promising for practical applications against Campylobacter coli and Campylobacter jejuni .