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SUMMARY: Campylobacter jejuni remains a leading cause of food-borne gastroenteritis worldwide, with contaminated poultry meat being a major transmission vector to humans. Effective control of C. jejuni colonisation in poultry is crucial for reducing public health risks. Among various intervention strategies, vaccination has emerged as a promising avenue. However, the journey towards a universally effective vaccine has not been met due variability in vaccine performance, and more research is still onging across the different species. Our comprehensive review critically examines the progress and problems in developing vaccines against C. jejuni. We concluded that regardless of the approaches used to develop vaccines, previously developed vaccines have effectively induced immunity but there were inconsistency among the results for the reduction in colonisation. Overall, partial success has been achieved in developing C. jejuni vaccine against poultry. This discrepancy highlights the complexity of vaccine development, which must deal with C. jejuni's antigenic diversity and the multifaceted nature of host–pathogen interactions. Additionally, the limited knowledge of immune system of chicken has hindered a full understanding of vaccine effectiveness against diverse C. jejuni. Our review highlights the need for further research on the genetic and phenotypic diversity of C. jejuni and the explorate the novel vaccine technology for development of successful C. jejuni vaccine. Additionally, the advances in understanding of C. jejuni pathogenesis and immune responses offer optimism for future vaccine development, ultimately aiming to reduce Campylobacter-related illness and enhance food safety in poultry products. [ABSTRACT FROM AUTHOR]

Aims To determine the effects of swarming motility (SM) and multi-locus sequence types (MLST) on the main effect of virulence genotype of Escherichia coli through an embryos lethality assay between the 12th and 18th days of incubation. Methods and results We collected 58 E. coli isolates from asymptomatic commercial hens (n  = 42) and lesions of colibacillosis cases (n  = 16), then classified their virulence genotype as avirulent, moderately virulent, virulent-healthy, and virulent-colibacillosis categories by the presence of five virulence-associated genes (iroN, ompT, hlyF, iutA , and iss). These isolates were further classified as non-motile, motile, or hyper-motile by SM assay. From the 58 isolates, we selected 29 for ELA and determined their MLST. Each isolate was inoculated into 15 embryonated eggs through the allantoic cavity. We found the avirulent isolates reduced the relative embryo weight compared to virulent-colibacillosis and moderately virulent isolates (37.49 vs. 41.51 and 40.34%, P  = 0.03). Among the moderately virulent and virulent-colibacillosis categories, embryo lethality was lower when isolates were non-motile. Yolk retention was unaffected by virulence categories, motility, or MLST. Conclusion Interaction between virulence genotype and SM substantially influenced the embryo lethality assay of E. coli isolates. [ABSTRACT FROM AUTHOR]

The obligate human pathogen Haemophilus influenzae utilizes a siderophore-independent (free) [Fe.sup.3+] transport system to obtain this essential element from the host iron-binding protein transferrin. The hFbpABC transporter is a binding protein-dependent ABC transporter that functions to shuttle (free) [Fe.sup.3+] through the periplasm and across the inner membrane of H. influenzae. This investigation focuses on the structure and function of the bFbpB membrane permease component of the transporter, a protein that has eluded prior characterization. Based on multiple-sequence alignments between permease orthologs, a series of site-directed mutations targeted at residues within the two conserved permease motifs were generated. The hFbpABC transporter was expressed in a siderophore-deficient Escherichia coli background, and effects of mutations were analyzed using growth rescue and radiolabeled [sup.55][Fe.sup.3+] transport assays. Results demonstrate that mutation of the invariant glycine (G418A) within motif 2 led to attenuated transport activity, while mutation of the invariant glycine (G155A/V/E) within motif 1 had no discernible effect on activity. Individual mutations of well-conserved leucines (L154D and LAI7D) led to attenuated and null transport activities, respectively. As a complement to site-directed methods, a mutant screen based on resistance to the toxic iron analog gallium, an bFbpABC inhibitor, was devised. The screen led to the identification of several significant hFbpB mutations; V497I, I174F, and $475I led to null transport activities, while S146Y resulted in attenuated activity. Significant residues were mapped to a topological model of the hFbpB permease, and the implications of mutations are discussed in light of structural and functional data from related ABC transporters.

Pathogenic Haemophilus influenzae, Neisseria spp. (Neisseria gonorrhoeae and N. meningitidis), Serratia marcescens, and other gram-negative bacteria utilize a periplasm-to-cytosol FbpABC iron transporter. In this study, we investigated the H. influenzae FbpABC transporter in a siderophore-deficient Escherichia coli background to assess biochemical aspects of FbpABC transporter function. Using a radiolabeled [Fe.sup.3+] transport assay, we established an apparent [K.sub.m] = 0.9 [micro]M and [V.sub.max] = 1.8 pmol/[10.sup.7] cells/min for FbpABC-mediated transport. Complementation experiments showed that hFbpABC is dependent on the FbpA binding protein for transport. The ATPase inhibitor sodium orthovanadate demonstrated dose-dependent inhibition of FbpABC transport, while the protonmotive-force-inhibitor carbonyl cyanide m-chlorophenyl hydrazone had no effect. Metal competition experiments demonstrated that the transporter has high specificity for [Fe.sup.3+] and selectivity for trivalent metals, including [Ga.sup.3+] and [Al.sup.3+], over divalent metals. Metal sensitivity experiments showed that several divalent metals, including copper, nickel, and zinc, exhibited general toxicity towards E. coli. Significantly, gallium-induced toxicity was specific only to E. coli expressing FbpABC. A single-amino-acid mutation in the gene encoding the periplasmic binding protein, FbpA(Y196I), resulted in a greatly diminished iron binding affinity [K.sub.d] = 5.2 x [10.sup.-4] [M.sup.-1], ~14 orders of magnitude weaker than that of the wild-type protein. Surprisingly, the mutant transporter [FbpA(Y196I)BC] exhibited substantial transport activity, ~35% of wild-type transport, with [K.sub.m] = 1.2 [micro]M and [V.sub.max] = 0.5 pmol/[10.sup.7] cells/min. We conclude that the FbpABC complexes possess basic characteristics representative of the family of bacterial binding protein-dependent ABC transporters. However, the specificity and high-affinity binding characteristics suggest that the FbpABC transporters function as specialized transporters satisfying the strict chemical requirements of ferric iron ([Fe.sup.3+]) binding and membrane transport.

The DNA sequence after the well-characterized Neisseria gonorrhoeae fbp gene has two open reading frames. One is called fbpB which encodes a protein which may act as a cytoplasmic permease and the other is called fbpC which encodes a protein which may act as a nucleotide-binding protein. The fbpABC operon composes an iron transport system that is homologous to the sfu and hit operons of Serratia marcescens and Haemophilus influenzae, respectively, and exhibits elements characteristic of ATP-binding cassette transporters.

The kinetics and mechanisms involved with loading of iron(III) into iron-free ferric binding protein A (FbpA) using iron(III) citrate as an iron source in the presence of excess citrate or phosphate (physiologically available anions) at pH 6.5 is reported. The findings demonstrated the important function of FbpA as a carrier for iron chelates as well as 'naked' or free iron acquisition processes of pathogenic Neisseria gonorrheae, Neisseria meningitidis, and Haemophilus influenzae.

The stability of unpurified proteins from rates of H/D exchange (SUPREX) is used for determining the identification and distribution of the synergistic anion in FeFbpA-X species in periplasmic preparations from Gram-negative bacteria. The studies have determined the identity of the in vivo anion bound to FeFbpA-X in the periplasm and has shown that the synergistic anion has played a structural and functional role in FbpA-mediated transport of iron across the periplasm and into the cytosol.

The obligate human pathogens Haemophilus influenzae, Neisseria gonorrhoeae, and N. meningitidis utilize a highly conserved, three-protein ATP-binding cassette transporter (FbpABC) to shuttle free Fe3+ from the periplasm and across the cytoplasmic membrane. The periplasmic binding protein, ferric binding protein (FbpA), is capable of transporting other trivalent cations, including Ga3+, which, unlike Fe3+, is not redox-active. Because of a similar size and charge as Fe3+, Ga3+ is widely used as a non-redox-active Fe3+ substitute for studying metal complexation in proteins and bacterial populations. The investigations reported here elucidate the similarities and differences in FbpA sequestration of Ga3+ and Fe3+, focusing on metal selectivity and the resulting transport function. The thermodynamic binding constant for Ga3+ complexed with FbpA at pH 6.5, in 50 mM 4-morpholineethanesulfonic acid, 200 mM KCl, 5 mM KH2PO4 was determined by UV-difference spectroscopy as $$ \log \,K_{{\text{eff}}}^\prime = 13.7 \pm 0.6. $$ This represents a 105-fold weaker binding relative to Fe3+ at identical conditions. The unfolding/refolding behavior of Ga3+ and Fe3+ holo-FbpA were also studied using a matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy technique, stability of unpurified proteins from rates of H/D exchange (SUPREX). This analysis indicates significant differences between Fe3+ and Ga3+ sequestration with regard to protein folding behavior. A series of kinetic experiments established the lability of the Ga3+FbpA–PO4 assembly, and the similarities/differences of stepwise loading of Fe3+ into apo- or Ga3+-loaded FbpA. These biophysical characterization data are used to interpret FbpA-mediated Ga3+ transport and toxicity in cell culture studies. [ABSTRACT FROM AUTHOR]

The obligate human pathogen Haemophilus influenzae utilizes a siderophore-independent (free) Fe3+ transport system to obtain this essential element from the host iron-binding protein transferrin. The hFbpABC transporter is a binding protein-dependent ABC transporter that functions to shuttle (free) Fe3+ through the periplasm and across the inner membrane of H. influenzae. This investigation focuses on the structure and function of the hFbpB membrane permease component of the transporter, a protein that has eluded prior characterization. Based on multiple-sequence alignments between permease orthologs, a series of site-directed mutations targeted at residues within the two conserved permease motifs were generated. The hFbpABC transporter was expressed in a siderophore-deficient Escherichia coli background, and effects of mutations were analyzed using growth rescue and radiolabeled 55Fe3+ transport assays. Results demonstrate that mutation of the invariant glycine (G418A) within motif 2 led to attenuated transport activity, while mutation of the invariant glycine (G155A/V/E) within motif 1 had no discernible effect on activity. Individual mutations of well-conserved leucines (L154D and L417D) led to attenuated and null transport activities, respectively. As a complement to site-directed methods, a mutant screen based on resistance to the toxic iron analog gallium, an hFbpABC inhibitor, was devised. The screen led to the identification of several significant hFbpB mutations; V497I, I174F, and S475I led to null transport activities, while S146Y resulted in attenuated activity. Significant residues were mapped to a topological model of the hFbpB permease, and the implications of mutations are discussed in light of structural and functional data from related ABC transporters. [ABSTRACT FROM AUTHOR]

We report the whole-genome sequences of Escherichia coli strains APEC-O2-MS1266 and APEC-O2-MS1657 isolated from the liver and heart of infected broilers in Mississippi State, US. The genomic information of these two causative strains may provide a valuable reference for comparative studies of avian pathogenic E. coli ., Competing Interests: The authors declare no conflict of interest.