Comparison of Two Culture-Based Detection Systems for the Isolation of Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii in Raw Ground Poultry.

Arcobacters are emerging pathogens that have been underestimated due to a lack of a standardized isolation method. The aim of this research was to evaluate the ability to isolate Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii using two Arcobacter-specific culture detection systems: (i) the Houf broth and modified charcoal cefoperazone deoxycholate agar supplemented with cefoperazone, amphotericin B, and teicoplanin (HB/mCCDA+CAT), and (ii) the Nguyen-Restaino-Juárez Arcobacter enrichment broth and chromogenic agar (NRJ-B/M). Both detection systems were evaluated for productivity ratio, sensitivity, and specificity. As a result, the productivity ratio for both plating agars were >90%, which indicates that the selective agents used in the two plating agars did not inhibit Arcobacter growth. Moreover, sensitivity evaluations using artificially inoculated retail ground poultry (n = 780) determined that both detection systems were able to isolate A. butlzeri with >95% sensitivity at the 0.1 and 1.0-2.0 CFU/g detection level. The sensitivity in A. cryaerophilus isolation was higher for NRJ-B/M (78.0% at 0.1 CFU/g; 95.1% at 1.0-2.0 CFU/g) when compared with HB/mCCDA+CAT (34.1% at 0.1 CFU/g; 51.2% at 1.0-2.0 CFU/g). Both detection systems resulted in <50% sensitivity when isolating A. skirrowii at 0.1 and 1.0-2.0 CFU/g; however, the sensitivity for NRJ-B/M was significantly higher than HB/mCCDA+CAT. At the detection level of 5.0 CFU/g, both detection systems were able to isolate A. skirrowii with 100% sensitivity. Specificity comparisons using uninoculated ground poultry samples (n = 40) indicated the growth of background microbiota were significantly inhibited or could be easily differentiated on NRJ-B/M (90.0%, specificity) when compared with HB/mCCDA+CAT (30.0%, specificity). Overall, these results show that the NRJ-B/M detection system is a more sensitive and specific detection system when isolating Arcobacter spp. from ground chicken.

NRJ Media as the Gold-Standard Arcobacter-Specific Detection System: Applications in Poultry Testing.

Arcobacter species are ubiquitous emerging pathogens with an impact that has been underestimated due to limitations in isolation and detection methods. Our group recently developed the novel NRJ Arcobacter-detection system, with major improvements in specificity and selectivity compared to other culture-based methods. In this work, the NRJ detection system was evaluated using retail whole broiler chicken carcass. Nanopore 16S rRNA gene amplicon sequencing demonstrated that Arcobacter species are found in very low abundance in retail chicken and that indigenous microbiota could be a major factor interfering with detection. Comparison of the microbiome obtained from modified Houf broth (HB) method, as the standard detection system, and the novel NRJ method, showed Arcobacter abundances of <15% and >97%, respectively. The NRJ system significantly inhibits the growth of non-target microbiota, and specifically allows the multiplication of Arcobacter species. In this report, we describe the gold-standard of Arcobacter-specific culture-based method to test food matrices, which can be used for other applications, such as clinical and environmental sampling.

A New Method for Detection of Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii Using a Novel Chromogenic Agar.

ABSTRACT: Arcobacter species are gram-negative rods that have been implicated in food- and waterborne illness. Although various cultural isolation methods have been proposed, the current procedures are unable to fully suppress the growth of background microbiota present in food samples, which inhibits Arcobacter isolation. The purpose of this study was to develop a selective enrichment broth and chromogenic plating medium to detect three Arcobacter species that have been recognized as emerging foodborne pathogens: Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii. The developed Nguyen-Restaino-Juárez (NRJ) Arcobacter detection system consists of a selective enrichment broth (NRJ-B) and a selective-differential plating medium (NRJ-M). The protocol of the detection method was determined by evaluating the growth of A. butzleri, A. cryaerophilus, and A. skirrowii under various temperatures (30, 35, and 42°C) and incubation (aerobic, microaerophilic, and anaerobic) conditions. Additionally, 47 Arcobacter strains and 39 non-Arcobacter strains were tested in inclusivity and exclusivity evaluations of NRJ-B and NRJ-M. Overall, the study determined that the optimal growth conditions of Arcobacter species using the NRJ Arcobacter detection system were aerobic incubation at 30°C. NRJ-B supported good growth of A. butzleri, A. cryaerophilus, and A. skirrowii while effectively suppressing the growth of non-Arcobacter strains after 48 h. Furthermore, NRJ-M yielded 97.8% inclusivity and 100.0% exclusivity using the tested strains and resulted in salmon-pigmented Arcobacter colonies (1.0 to 1.5 mm in diameter) after 72 h. The novel protocol is the first to develop a chromogenic plating medium for the isolation of Arcobacter species. This simple and accurate test method would greatly contribute to understanding the distribution of pathogenic Arcobacter species in food samples.

Dendritic spine abnormalities in amyloid precursor protein transgenic mice demonstrated by gene transfer and intravital multiphoton microscopy.

Accumulation of amyloid-beta (Abeta) into senile plaques in Alzheimer's disease (AD) is a hallmark neuropathological feature of the disorder, which likely contributes to alterations in neuronal structure and function. Recent work has revealed changes in neurite architecture associated with plaques and functional changes in cortical signaling in amyloid precursor protein (APP) expressing mouse models of AD. Here we developed a method using gene transfer techniques to introduce green fluorescent protein (GFP) into neurons, allowing the investigation of neuronal processes in the vicinity of plaques. Multiphoton imaging of GFP-labeled neurons in living Tg2576 APP mice revealed disrupted neurite trajectories and reductions in dendritic spine density compared with age-matched control mice. A profound deficit in spine density (approximately 50%) extends approximately 20 mum from plaque edges. Importantly, a robust decrement (approximately 25%) also occurs on dendrites not associated with plaques, suggesting widespread loss of postsynaptic apparatus. Plaques and dendrites remained stable over the course of weeks of imaging. Postmortem analysis of axonal immunostaining and colocalization of synaptophysin and postsynaptic density 95 protein staining around plaques indicate a parallel loss of presynaptic and postsynaptic partners. These results show considerable changes in dendrites and dendritic spines in APP transgenic mice, demonstrating a dramatic synaptotoxic effect of dense-cored plaques. Decreased spine density will likely contribute to altered neural system function and behavioral impairments observed in Tg2576 mice.