Salmonellosis in Poultry Flocks: Pathogenesis, Rapid Detection, and Food Safety Implications

Introduction

Salmonellosis remains one of the most economically burdensome bacterial diseases in commercial poultry production worldwide. The disease is caused by members of the genus Salmonella enterica subsp. enterica, with serovars Enteritidis and Typhimurium representing the predominant zoonotic threats associated with egg and meat consumption [1, 2]. Poultry flocks serve as primary reservoirs, and contamination can occur at any stage from hatchery to slaughter [3]. The public health impact is substantial: human salmonellosis cases attributable to poultry have not declined in proportion to reductions in carcass contamination, indicating that virulence attributes of persistent serovars and the limitations of conventional detection warrant continued investigation [4, 5].

This review provides an exhaustive examination of the pathogenetic mechanisms of Salmonella in poultry, modern rapid detection platforms, and integrated food safety measures spanning preharvest through postharvest interventions.

Pathogenesis of Salmonella in Poultry

Bacterial Virulence Factors

The pathogenic potential of non-typhoidal Salmonella serovars in poultry is largely determined by genes encoded within Salmonella Pathogenicity Islands (SPIs). SPI-1 and SPI-2 encode type III secretion systems (T3SS-1 and T3SS-2) that inject effector proteins into host enterocytes, facilitating invasion and intracellular survival [1]. SPI-1 effectors such as SopE, SopB, and SipA induce membrane ruffling and bacterial uptake, while SPI-2 effectors enable replication within Salmonella-containing vacuoles. Additional virulence determinants include flagella (FliC), fimbriae (e.g., Fim, Lpf, Bcf), and lipopolysaccharide (LPS) O antigens that contribute to adhesion, biofilm formation, and immune evasion [6, 1].

Serovar-specific variation in virulence gene repertoires influences the clinical outcome. S. Enteritidis possesses the sef operon encoding SEF14 fimbriae, which enhances colonization of reproductive tissues and vertical transmission. S. Typhimurium relies on the fliC flagellin gene and a distinct set of effectors that promote invasion of the cecal epithelium [7, 8].

Infection Routes and Clinical Manifestations

Salmonella can be transmitted horizontally through contaminated feed, water, litter, and fomites, and vertically via transovarian infection from infected breeder flocks [9, 1, 3]. The hatchery is a critical point of amplification: meta-analysis indicates that hatchery-related sources contribute a 48.5% prevalence of Salmonella positivity in broilers [3]. Ingested bacteria traverse the proventriculus and gizzard, colonize the ceca and ileum, and may disseminate to the spleen, liver, and oviduct.

Clinical signs vary with age, serovar, and immune status. In chicks, S. Typhimurium and S. Enteritidis produce diarrhea, lethargy, omphalitis, and increased mortality. Older birds often become asymptomatic carriers, shedding bacteria intermittently in feces [10, 2]. Carrier birds pose a substantial risk for flock-to-flock transmission and contamination of eggs and carcasses at processing [11].

Host Immune Response and Carrier State

The avian immune system responds to Salmonella through both innate and adaptive arms. Macrophages and heterophils are recruited to the site of infection, and T-cell mediated immunity is essential for clearance [1]. Humoral responses generate anti-LPS and anti-flagellin antibodies, which are exploited for serological monitoring via Enzyme-Linked Immunosorbent Assay (ELISA) (see Enzyme-Linked Immunosorbent Assay (ELISA) for Feline Leukemia Virus for analogous methods). However, Salmonella can evade host defenses by persisting within macrophages, leading to a prolonged carrier state that is difficult to detect by culture [12, 11].

Rapid Detection of Salmonella in Poultry Flocks

Conventional Culture and Its Limitations

Traditional isolation involves pre-enrichment in buffered peptone water, selective enrichment (e.g., Rappaport-Vassiliadis broth), and plating on selective agars such as xylose lysine deoxycholate (XLD) agar, where Salmonella produces characteristic black-centered colonies [10, 8]. These methods require 5 to 7 days and lack sensitivity for low-level shedding or stressed cells. This delay impedes timely intervention.

Molecular Detection Methods

Real-Time PCR (qPCR)

Real-time PCR targeting the invA gene, which is conserved across Salmonella serovars, provides rapid and specific identification directly from fecal or environmental samples. The assay amplifies a 423 bp fragment of invA with high sensitivity, detecting as few as 10^2 CFU/g [10, 8]. Multiplex qPCR formats can simultaneously differentiate serogroups by targeting serovar-specific genes: prot6 for S. Enteritidis and fliC for S. Typhimurium [8].

Quantitative PCR combined with most probable number (MPN) enumeration yields absolute counts, enabling risk stratification based on bacterial load [5]. A framework using combined quantification and deep serotyping (e.g., CRISPR-SeroSeq) has been developed to generate risk scores for broiler flocks, integrating the presence of key performance indicator (KPI) serovars (Enteritidis, Infantis, Typhimurium) with total Salmonella quantity [5].

Whole Genome Sequencing (WGS)

WGS provides the highest resolution for epidemiological tracing, antimicrobial resistance (AMR) profiling, and virulence gene characterization. By sequencing the entire bacterial genome, one can identify plasmid-mediated resistance determinants (e.g., blaCTX-M, tet(A), sul1) and clonal relationships between poultry and human isolates [13]. Multilocus sequence typing (MLST) derived from WGS data has revealed the international dissemination of multidrug-resistant S. Infantis ST32 clones carrying megaplasmids [13]. The application of WGS in routine surveillance is limited by cost and bioinformatics infrastructure, but its value for outbreak investigations is undisputed.

Mass Spectrometry and Alternative Assays

Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry enables identification of Salmonella from culture colonies based on ribosomal protein profiles. This technique offers rapid (minutes) and accurate identification to genus and serogroup level, though serovar resolution may require supplementary analysis. Loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA) represent field-deployable alternatives that require minimal instrumentation [14].

A decision tree summarizing the selection of detection methods based on flock status and objective is presented in Figure 1.

flowchart TD
    A[Flock: Clinical signs or risk], > B{Objective}
    B, >|Rapid screening| C[Pooled litter/boot swabs]
    C, > D[qPCR (invA)]
    D, > E{Positive?}
    E, >|No| F[No further action]
    E, >|Yes| G[Confirm serovar by multiplex PCR]
    G, > H[Option: WGS for AMR typing]
    B, >|Regulatory testing| I[Culture per standard protocols]
    I, > J[Biochemical confirmation]
    J, > K[Serotyping (antisera or PCR)]
    B, >|Epidemiological investigation| L[WGS with core genome MLST]
    L, > M[Compare with human isolates]

Figure 1. Decision workflow for Salmonella detection in poultry flocks.

Food Safety Implications and Farm-to-Fork Prevention

Preharvest Interventions

Biosecurity is the cornerstone of Salmonella control. Measures include implementing all-in/all-out production, cleaning and disinfecting houses between flocks, controlling vectors (rodents, insects, wild birds), and using dedicated footwear and equipment for each house [15, 9, 16]. A study of backyard flocks in France revealed that biosecurity compliance is heterogeneous: 50% of owners did not wash hands systematically, and over 60% did not use dedicated shoes [15]. Such gaps in small-scale operations represent a continuing food safety challenge.

Vaccination of breeder and layer flocks reduces shedding and vertical transmission. Live attenuated vaccines (e.g., S. Gallinarum 9R) and killed bacterins are available; the SG 9R vaccine has been shown to reduce clinical salmonellosis in breeder flocks in Sri Lanka [17, 11]. However, vaccine efficacy varies by serovar and must be combined with hygienic measures [11].

Feed additives including prebiotics, probiotics, postbiotics, and bacteriophages are being explored as alternatives to antimicrobial growth promoters [1, 18, 19]. Lactobacillus strains isolated from healthy ceca exhibit anti-Salmonella activity in vitro, suggesting probiotic potential for competitive exclusion [19]. Bacteriophage therapy has demonstrated success in reducing Salmonella colonization in challenged birds, but requires careful cocktail design to prevent resistance emergence [18].

Processing and Postharvest Control

At the slaughterhouse, interventions such as carcass washing with organic acids (e.g., lactic acid), chlorinated water, and hot water sprays reduce surface contamination. However, no single "kill step" can eliminate Salmonella from raw poultry without compromising product quality [14]. Risk management options must consider contamination levels entering the plant; preharvest reduction remains the most effective strategy.

Deep serotyping and quantification of Salmonella entering the plant allow processors to allocate higher-risk flocks to enhanced interventions (e.g., extended chilling, irradiation) [5]. Regulatory frameworks such as those in the European Union target specific serovars (Enteritidis, Typhimurium, Infantis) for mandatory control in breeding and laying flocks [4, 16].

Antimicrobial Resistance as a Food Safety Threat

Multidrug-resistant (MDR) Salmonella serovars, especially S. Infantis, have emerged globally. In Korea, MDR S. Infantis strains isolated from broiler flocks between 2020 and 2023 harbored megaplasmids carrying resistance genes to ampicillin, cephalosporins, chloramphenicol, nalidixic acid, and tetracycline [13]. Such MDR strains can persist in the food chain and cause human infections that are difficult to treat. Surveillance of AMR trends is essential for informing treatment guidelines and intervention priorities [20].

Conclusion

Salmonellosis in poultry is a multifactorial disease that demands integrated control strategies targeting every link in the production chain. Advances in rapid molecular diagnostics, particularly real-time PCR and whole genome sequencing, have enhanced our ability to detect low-level contamination and track the dissemination of virulent, drug-resistant clones. These tools, when coupled with rigorous biosecurity, vaccination, and rational antimicrobial use, hold the potential to reduce the burden of poultry-associated human salmonellosis. Continued research into host-pathogen interactions, alternative therapies, and risk-based food safety management will be essential to protect both animal health and public health [1, 21, 22].

References

[1] Shaji S, Selvaraj R, Shanmugasundaram R. Salmonella Infection in Poultry: A Review on the Pathogen and Control Strategies. Microorganisms. 2023. https://www.semanticscholar.org/paper/09626066cb93dcefde5fff585f2a643ae8662dc4

[2] Tariq S, Samad A, Hamza M, et al. Salmonella in Poultry; An Overview. International Journal of Multidisciplinary Sciences and Arts. 2022. https://www.semanticscholar.org/paper/36c572f0a409bc6b54ca20ba0095cbac83ec824f

[3] Wang J, Vaddu S, Bhumanapalli S, et al. A systematic review and meta-analysis of the sources of Salmonella in poultry production (pre-harvest) and their relative contributions to the microbial risk of poultry meat. Poultry Science. 2023. https://www.semanticscholar.org/paper/95ea413a335b25d8855d37a26a63782cba917083

[4] Koutsoumanis K, Allende A, Álvarez-Ordoñez A, et al. Salmonella control in poultry flocks and its public health impact. EFSA Journal. 2019. https://www.semanticscholar.org/paper/5a5a418da49ad91b80bac5d3ad9b5b5748f44eed

[5] Obe T, Siceloff AT, Crowe MG, et al. Combined Quantification and Deep Serotyping for Salmonella Risk Profiling in Broiler Flocks. Applied and Environmental Microbiology. 2023. https://www.semanticscholar.org/paper/d7e75397f70e1c63cae15fae8698e04e9944eb7e

[6] Dar M, Ahmad SM, Bhat S, et al. Salmonella typhimurium in poultry: a review. Journal. 2017. https://www.semanticscholar.org/paper/40827a1ae7fec715dabd0246e54e3f993dcd981d

[7] De Van Giessen WA, et al. The identification of Salmonella enteritidis-infected poultry flocks associated with an outbreak of human salmonellosis. Epidemiology and Infection. 1992. https://www.semanticscholar.org/paper/1e6f2d90cc2a4f1a7d5edff14abd2c092fe05008

[8] Afshari A, Baratpour A, Khanzade S, et al. Salmonella Enteritidis and Salmonella Typhimorium identification in poultry carcasses. Iranian Journal of Microbiology. 2018. https://www.semanticscholar.org/paper/52808301af14f2456a4513bf95f06d58c4ada981

[9] Naumovska S, Vukomanović AG, Pajić M, et al. Biosecurity Measures for Prevention and Control of Salmonella in Poultry Farms. Contemporary Agriculture. 2025. https://www.semanticscholar.org/paper/6eda26d641da4fd61bcc9e1f4d310f5f9f2a380b

[10] Sonkusale PM, Warke SR, Chaitanya S. Pathological Studies and Molecular Detection of Salmonellosis in Desi Chickens of Nagpur Region. International Journal of Research and Review. 2023. https://www.semanticscholar.org/paper/5b780ea9722bd757e93ffa36037f18dad4ae0ea4

[11] Liyanagunawardena N, Fernando P, Madalagama R, et al. Control of Poultry Salmonellosis - Detection of Carrier Status in Grandparent Farms in Sri Lanka. Journal. 2016. https://www.semanticscholar.org/paper/c638e5c6c18d397203719ca8af3d8706147a088c

[12] Sahu R, Patil S, Lalsangzuala C. Salmonellosis in poultry - an overview. Journal. 2015. https://www.semanticscholar.org/paper/f0ec9c8d0a694821e3853fab13b145319b3fff91

[13] Lee SH, Lee OM, Kang SI, et al. Recent Occurrence and Rapid Spread of Multidrug-Resistant Salmonella Infantis in Broiler Flocks in Korea. Foodborne Pathogens and Disease. 2025. https://www.semanticscholar.org/paper/ada1d7485f4db27c536fe8597e1377d604b2feea

[14] Cohn A. Risk Management Options to Reduce Human Salmonellosis Cases Due to Consumption of Raw Poultry. Food Protection Trends. 2023. https://www.semanticscholar.org/paper/3f1e0f66f6c4c3c77e36b03f48d208b8b5cbed02

[15] Souvestre M, Delpont M, Guinat C, et al. Backyard poultry flocks in France: A diversity of owners and biosecurity practices. Preventive Veterinary Medicine. 2021. https://www.semanticscholar.org/paper/15b01cf499c87666a94cc038560344e9ef165d45

[16] Huneau-Salaün A, Le Bouquin S, Chemaly M. Salmonella Control Programme in France: Factors Influencing the Detection of Salmonella in Laying Hen Flocks From 2013 to 2021. Zoonoses and Public Health. 2025. https://www.semanticscholar.org/paper/cb2d90942ab1ccf6f4cf93c94ed4fef142904291

[17] Liyanagunawardena N, Fernando P, Weerasooriya G, et al. SG 9R Vaccine to Control Salmonellosis in Poultry Breeder Flocks in Sri Lanka. Journal. 2016. https://www.semanticscholar.org/paper/547e87ca593e6d7af3d13907b11bc263455a0cf3

[18] Wernicki A, Nowaczek A, Urban-Chmiel R. Bacteriophage therapy to combat bacterial infections in poultry. Virology Journal. 2017. https://www.semanticscholar.org/paper/215d3f3d20a4ade3d6bfca6ec7260cfad8f06264

[19] Chuwatthanakhajorn S, Wedley A, Watts A, et al. Lactobacilli Isolated from the Caecum of Healthy Broilers with Antimicrobial Activity are Probiotic Candidates for Controlling Salmonella. Probiotics and Antimicrobial Proteins. 2026. https://pubmed.ncbi.nlm.nih.gov/41855006/

[20] Hossain MJ, Attia Y, Ballah F, et al. Zoonotic Significance and Antimicrobial Resistance in Salmonella in Poultry in Bangladesh for the Period of 2011–2021. Zoonotic Diseases. 2021. https://www.semanticscholar.org/paper/1bf616245ebb00013959e386bf2c678401058b9e

[21] Ayala Velastegui D, Shariat NW. Investigating the differences in Salmonella serovar transmission within broiler production. Frontiers in Veterinary Science. 2026. https://pubmed.ncbi.nlm.nih.gov/42078842/

[22] Cason EE, Rivera MP, Maradiaga S, et al. Temporal Sampling of Commercial Turkey Flocks Highlights Challenges of Preharvest Salmonella enterica Monitoring. Journal of Food Protection. 2026. https://pubmed.ncbi.nlm.nih.gov/41956287/

[23] Barton Behravesh C, Brinson D, Hopkins BA, et al. Backyard Poultry Flocks and Salmonellosis: A Recurring, Yet Preventable Public Health Challenge. Clinical Infectious Diseases. 2014. https://www.semanticscholar.org/paper/72393cdea21cac7d5ac82f4a800b4d08753d7923

[24] Hosseinzadeh S, Saei HD, Ahmadi M, et al. Antimicrobial effect of Licochalcone A and Epigallocatechin-3-gallate against Salmonella Typhimurium isolated from poultry flocks. Iranian Journal of Microbiology. 2018. https://www.semanticscholar.org/paper/09886534f5b2325311a2d5331a5d45b1a31478d6

[25] Basler C. Changing Trends in Backyard Flocks: Review of Outbreaks of Human Salmonellosis Linked to Contact with Live Poultry, United States, 1990-2013. Journal. 2014. https://www.semanticscholar.org/paper/d06d44eadae4ef2f1bb1930967d23eb06c43f722

[26] Ansari F, Pourjafar H, Bokaie S, et al. Association between Poultry Density and Salmonella Infection in Commercial Laying Flocks in Iran using a Kernel Density. Journal. 2017. https://www.semanticscholar.org/paper/483b03cc7c1c8d75f32f2391bd02e9894f0d04de

[27] Beninca ALV, Candeias A, Fernandes SR, et al. Efficiency of sanitary management of litter in Eimeria spp. control in poultry housing of Western Paraná, Brazil. Brazilian Journal of Veterinary Parasitology. 2021. https://www.semanticscholar.org/paper/240c4198aa51031684eb8997580892479df5934d

[28] Reilly W, Forbes GI, Sharp J, et al. Poultry-borne salmonellosis in Scotland. Epidemiology and Infection. 1988. https://www.semanticscholar.org/paper/8ab6111e9f19959b2dc8c4e9106e056351d1e6ce

[29] Ager EO, Nickodem CA, Brown J, et al. Diet-vaccine interactions: SQM Iron and Salmonella vaccination shape poultry gut microbiota. Applied and Environmental Microbiology. 2026. https://pubmed.ncbi.nlm.nih.gov/41979594/

[30] Bertin B, Bayon-Auboyer MH, Fellag M, et al. Reliable Differentiation of a Bivalent Live Salmonella Vaccine and Field Strains: Multi-Supplier Validation of a Disc Diffusion Method. Veterinary Sciences. 2026. https://pubmed.ncbi.nlm.nih.gov/41893720/