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Segmented filamentous bacteria-based treatment to elicit protection against Enterobacteriaceae in Layer chickens - PubMed

  • ️Sun Jan 01 2023

Segmented filamentous bacteria-based treatment to elicit protection against Enterobacteriaceae in Layer chickens

Jared Meinen-Jochum et al. Front Microbiol. 2023.

Abstract

Introduction: Gut microbes like segmented filamentous bacteria (SFB) play a key role in gut maturation during early life, as demonstrated in humans and mice. Our previous study demonstrated oral inoculation of ileum-spores containing SFB to chickens after hatch increases early SFB gut colonization, which increases immune maturation and resistance to bacteria, like Salmonella, as tested in vitro; however, more studies are needed for treatment optimization and in vivo testing. The objectives of this study were to (1) test a treatment that includes both spores and filamentous SFB, (2) validate antimicrobial ability of the treatment in layer hens in vivo, and (3) elucidate its molecular mechanism.

Methods: One-day-old specific pathogen-free layers (n = 12 per group) were orally treated with either PBS (CON) or SFB-based treatment (SFB). At 4 days post-inoculation (DPI), both CON and SFB groups were orally challenged with Salmonella Typhimurium. Total Enterobacteriaceae and Salmonella were examined by plating and enumeration in feces at 7,10 and 14 dpi; and in the ileum, cecum, and spleen at 16 dpi in euthanized birds. The presence and levels of SFB were determined from ilea scrapings via microscopy and qPCR, respectively. Relative gene expression of host-derived antimicrobial peptides and cytokines in the distal ileum was determined by RT-qPCR.

Results: At 10 and 14 dpi, a significant decrease in total Enterobacteriaceae was observed in the feces of the SFB group. At necropsy, the level of SFB was significantly higher in the SFB group than in the CON group, while a significant decrease in total Enterobacteriaceae and Salmonella was observed in the ceca of the SFB group. RT-qPCR revealed increased expression of β-defensin 14, and cytokines IL-10 and IFNγ.

Discussion: The introduction of SFB at hatch as a prophylactic treatment may benefit commercial partners as well as consumers by reducing the incidence of Enterobacteriaceae in food animals. Reduction of these bacteria in animals would, in turn, increase animal health, productivity, and safety for consumers. Studies to optimize the treatment for poultry industry applications are ongoing in our lab.

Keywords: Enterobacteriaceae; antimicrobial peptides; cytokines; layer chickens; segmented filamentous bacteria.

Copyright © 2023 Meinen-Jochum, Ott and Mellata.

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Conflict of interest statement

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.

Figures

Figure 1
Figure 1

Detection of Segmented Filamentous Bacteria (SFB) via Gram-Staining and FISH Microscopy in the Pilot Inoculation Experiment. Representative images of ilea scrapings analyzed by gram-stain (A) and FISH microscopy (B) demonstrate the inability to detect SFB filaments in Control birds prior to 16-days post inoculation (dpi) and presence of filaments in SFB treated birds after 5 dpi. Arrows indicate intracellular offspring (blue) or filamentous SFB (white).

Figure 2
Figure 2

Ilea-associated Segmented Filamentous Bacteria (SFB) Quantification. Log10 CFU/g of SFB detected in ilea scrapings calculated by qPCR in (A) the pilot inoculation experiment at respective 2-, 5-, and 16- days post inoculation (dpi) and (B) Salmonella challenge study at final necropsy (16 dpi). ****, p < 0.0001. Dots indicate individual birds. The horizontal dashed line represents the limit of detection (LOD). Samples that did not demonstrate amplification in qPCR assays were set to half of the LOD.

Figure 3
Figure 3

Detection of Segmented Filamentous Bacteria (SFB) via Gram-Staining and FISH Microscopy in Salmonella Challenge Study. Representative images of ilea scrapings analyzed by gram-stain (A) and FISH microscopy (B) demonstrate the inability to detect SFB in Control birds and presence of SFB in SFB treated birds at final necropsy (16-days post inoculation). White arrows indicate filamentous SFB.

Figure 4
Figure 4

Fecal Tracking of Total Enterobacteriaceae. Total Enterobacteriaceae in feces in the pilot inoculation experiment at 11- and 14-days post inoculation (dpi) (A) and in the Salmonella challenge at 7-, 10-, and 14-dpi (B). *, p < 0.05; **, p < 0.01. Each dot represents a separate bird tested.

Figure 5
Figure 5

Salmonella Enumeration at Final Necropsy. Total Salmonella enumerated in the (A) ceca (B) spleen and (C) ileum at final necropsy, 16-days post inoculation. ***, p < 0.001. Dots indicate individual birds. The horizontal dashed line represents the limit of detection (LOD). Samples that remained negative after enrichment were assigned a 0 value.

Figure 6
Figure 6

Total Enterobacteriaceae Enumeration at Final Necropsy. Total Enterobacteriaceae enumerated in the ceca (A), spleen (B), and ileum (C) at 16-days post inoculation (dpi). **, p < 0.01. Dots indicate individual birds. The horizontal dashed line represents the limit of detection (LOD). Samples that remained negative after enrichment were assigned a 0 value.

Figure 7
Figure 7

Changes in Antimicrobial Peptide and Interleukin Gene Expression. Log2 fold change in gene expression of select antimicrobial peptides (AMPs) (A) and interleukins (B). *, p < 0.05; **, p < 0. 01; ****, p < 0.0001. ND denotes no amplification detected.

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