Antibiotic Resistance Patterns of Major Zoonotic Pathogens from All-Natural, Antibiotic-Free, Pasture-Raised Broiler Flocks in the Southeastern United States.

The use of antibiotics in agroecosystems has been implicated in the rise in antibiotic resistance (AR), which can affect environmental, animal, and human health. To determine the environmental impact of antibiotic use in agroecosystems, appropriate background levels of AR in agricultural environments in the absence of antibiotic application must be determined. Therefore, to determine background levels of AR in broiler production, four target microbes (, , , and ) were isolated from 15 all-natural, antibiotic-free, pasture-raised broiler flocks from six farms within the southeastern United States. The AR profiles of these isolates were characterized using the CDC National Antimicrobial Resistance Monitoring System for Enteric Bacteria (NARMS), and these resistance patterns were compared across target microbes and farms and throughout the life cycle of the flocks along the farm-to-fork continuum. Antibiotic resistances were most prevalent in and and least prevalent in . Although and were isolated from the same farms and characterized using the same NARMS plates, they exhibited distinct AR profiles, with demonstrating clear farm-specific resistance patterns. Multidrug resistance rates (three or more antibiotics), in order of prevalence, were (63.9%), (36.0%), (12.7%), and (1.4%). The results of this study demonstrate the variability in background AR among major food safety-related microbes, even when isolated from similar production and processing samples from the same farms, and indicate the need for the proper design of future broiler production studies to account for this highly dynamic background AR.

Salmonella Biomapping of a Commercial Broiler Hatchery.

Poultry-associated salmonellosis results in significant costs to poultry producers and consumers. Given the vertically integrated nature of the United States poultry industry, a better understanding of Salmonella ecology throughout all levels of poultry production is essential. One nexus point is the hatchery, where eggs from multiple broiler breeder farms are incubated and hatched, with the chicks being sent to numerous farms; therefore, the hatchery represents an ideal area to understand preharvest Salmonella ecology and flow. To achieve this, a commercial broiler hatchery was biomapped, focusing on Salmonella prevalence and serotype diversity among four major sample type categories (Air, Egg, Water, Facility) across five different places in the prehatch, hatch, and posthatch areas. Following two sets of eggs from broiler breeder farms over two production days, the overall Salmonella prevalence was 26% (48/184). Of the positive samples, the highest prevalence was observed in swabs taken from the floor drains in the facility and transport truck (56%), as well as in the hatch and posthatch hatchery areas (50%). Kentucky (n = 17), Gaminara (n = 12), and Alachua (n = 11) were the dominant Salmonella serotypes, with serotypes of greatest outbreak concern from chickens (Enteritidis) representing only 6.25% (3/48) of all recovered Salmonella isolates. The posthatch transport area, including the underfloor reservoirs of the transport trucks, not only harbored Enteritidis but also the enrichment broths from these Salmonella-positive samples also possessed sequences matching the commercial live-attenuated vaccine Typhimurium strain according to CRISPR SeroSeq analyses. These findings highlight the complex diversity of commercial hatchery Salmonella populations, including identifying facility floor drains and transport trucks as potentially important critical control points for hatchery managers to focus their Salmonella mitigation efforts to reduce loads and serotypes entering live production farms.

16S amplicon-based microbiome biomapping of a commercial broiler hatchery.

Hatcheries, where eggs from multiple breeder farms are incubated and hatched before being sent to different broiler farms, represent a nexus point in the commercial production of broilers in the United States. Considering all downstream microbial quality and safety aspects of broiler production (live production, processing, consumer use) can be potentially affected by the hatchery, a better understanding of microbial ecology within commercial hatcheries is essential. Therefore, a commercial broiler hatchery was biomapped using 16S rRNA amplicon-based microbiome analyses of four sample type categories (Air, Egg, Water, Facility) across five different places in the pre-hatch, hatch, and post-hatch areas. While distinct microbiota were found for each sample type category and hatchery area, microbial community analyses revealed that Egg microbiota trended towards clustering with the facility-related samples when moving from the prehatch to post-hatch areas, highlighting the potential effect of the hatchery environment in shaping the pre-harvest broiler-related microbiota. Prevalence analyses revealed 20 ASVs (Core20) present in the core microbiota of all sample types and areas, with each ASV possessing a unique distribution throughout the hatchery. Interestingly, three Enterobacteriaceae ASVs were in the Core20, including Salmonella. Subsequent analyses showed that Salmonella, while a minor prehatch and hatch Core20ASV, dominated the Enterobacteriaceae niche and total microbiota in the chick pad feces in the post-hatch area of the hatchery, and the presence of this Salmonella ASV in the post-hatch feces was associated with swabs of breakroom tables. These findings highlight the complexity of commercial hatchery microbiota, including identifying chick pad feces and breakroom tables as potentially important sampling or disinfection targets for hatchery managers to focus their Salmonella mitigation efforts to reduce loads entering live production farms.

Tissue Colonization and Egg and Environmental Contamination Associated with the Experimental Infection of Cage-Free Laying Hens with Salmonella Braenderup.

In 2018, a national recall of shell eggs in the United States occurred due to human illness caused by Salmonella Braenderup. Although previous studies have identified Salmonella Braenderup in laying hens and the production environment, little is known about the ability of this Salmonella serovar to infect laying hens and contaminate eggs. The objective of this study was to examine the invasiveness of Salmonella Braenderup in laying hens as well as its ability to persist in the production environment. Specific-pathogen-free laying hens (four trials; 72 hens/trial) were orally challenged with 107 colony-forming units of Salmonella Braenderup. On day 6 postinoculation, half of the challenged hens were euthanatized, and samples of ileocecal junction (sections above and below it, and portions of both ceca), liver, spleen, ovary, and oviduct tissues were collected and cultured for Salmonella Braenderup. Egg and environmental (nest box swaps and substrate (litter)) samples were collected days 7-20 postinoculation (Trials 1 and 2; excluding weekends) and days 7-27 postinoculation (Trials 3 and 4; excluding weekends) to detect Salmonella Braenderup. Recovery of Salmonella Braenderup was highest in ileocecal tissue samples (11.1%-33.3%; P < 0.05), with little to no recovery in other collected tissue samples. Salmonella Braenderup was detected in a small number of shell emulsions (0%-2.9%; P < 0.01) and recovered in Trial 1 at a high rate (92.5%; P < 0.0001) in the substrate composite samples; however, recovery of Salmonella Braenderup was low in the other egg and environmental samples. These trials indicate that Salmonella Braenderup is not an invasive Salmonella serovar for cage-free laying hens, especially when compared to serovars of concern to the egg industry. However, it may persist in the environment at low levels.

Colonización de tejidos y contaminación ambiental y de huevo asociados con la infección experimental de gallinas de postura libres de jaulas por Salmonella Braenderup. En 2018, se retiraron del mercado a nivel nacional en los Estados Unidos huevos con cascarón debido a una enfermedad en humanos causada por Salmonella Braenderup. Aunque estudios anteriores han identificado Salmonella Braenderup en gallinas de postura y en ambientes de producción, se conoce poco sobre la capacidad de esta serovariedad de Salmonella para infectar a las gallinas ponedoras y contaminar el huevo. El objetivo de este estudio fue examinar la capacidad de invasión de Salmonella Braenderup en gallinas ponedoras, así como su capacidad para persistir en el ambiente de producción. Se desafiaron oralmente a gallinas de postura libres de patógenos específicos (cuatro ensayos; 72 gallinas/ensayo) con 107 unidades formadoras de colonias de Salmonella Braenderup. El día seis después de la inoculación, la mitad de las gallinas desafiadas se sacrificaron y se recolectaron y cultivaron muestras de la unión ileocecal (secciones anteriores y posteriores de la misma y porciones de ambos ciegos), hígado, bazo, ovario y oviducto y se cultivaron para Salmonella Braenderup. Se recolectaron muestras de huevos y ambientales (hisopos de las cajas de nido y sustrato [cama] en los días 7 a 20 después de la inoculación (Pruebas 1 y 2; excluyendo los fines de semana) y en los días 7 a 27 después de la inoculación (Pruebas 3 y 4; excluyendo los fines de semana) para detectar Salmonella Braenderup. La recuperación de Salmonella Braenderup fue mayor en las muestras de tejido ileocecal (11.1%­33.3%; P < 0.05), con poca o ninguna recuperación en otras muestras de tejido recolectadas. Se detectó Salmonella Braenderup en un pequeño número de emulsiones de cascarones (0%­2.9%; P < 0.01) y se recuperó en el Ensayo 1 a una tasa alta (92.5%; P < 0.0001) en las muestras compuestas de sustrato; sin embargo, la recuperación de Salmonella Braenderup fue baja en las otras muestras de huevos y ambientales. Estos ensayos indican que Salmonella Braenderup no es un serovar de Salmonella invasivo para gallinas de postura sin jaulas, especialmente cuando se compara con los serovares de interés para la industria del huevo. Sin embargo, puede persistir en el medio ambiente en niveles bajos.