Recipient Organization
PENNSYLVANIA STATE UNIVERSITY
408 Old Main
UNIVERSITY PARK,PA 16802-1505
Performing Department
(N/A)
Non Technical Summary
Salmonella enterica serotype Dublin is adapted to cattle and can also infect humans. It can persist on farms for months to years and can be extremely difficult to treat due to its multi-drug resistance. This strain is slowly increasing in prevalence across the United States and can result insevere economic loss. However, no data are available regarding the herd prevalence of S. Dublin in Pennsylvania dairy herds, despite this pathogen being isolated from several human cases in Pennsylvania since 2014, and many animal cases in the past two years.This project aims to address the potentially underdiagnosed problems caused by Salmonella Dublin to the Pennsylvania dairy industry. We will screen bulk tank milk samples for S. Dublin antibodies to determine how widespread it is. For positive herds, we will work with farmers and use advanced detection methods to isolate the bacteria from animals and the environment in a "seek and destroy" approach. We will also use a structured risk assessment tool to evaluate farms and identify major points of transmission to create a tailored mitigation approach designed based on risk assessment in each herd.We will develop a suite of extension resources to increase the level of knowledge aboutSalmonellaDublin, and to change attitudes and behaviors toward the prevention, detection, and control ofSalmonellain dairy herds. We will leverage the extensive reach of Penn State Extension to provide educational resources for PA dairy farm owners and employees, as well as the veterinarians who work with these dairy farms. These resources and engagement opportunities will increase knowledge and positively impact attitudes and behaviors about preventing and controllingS. Dublin in dairy herds and reducing the risk of zoonotic disease, ultimately leading to improved management and biosecurity inS.Dublin positive herds in PA.
Animal Health Component
100%
Research Effort Categories
Basic
(N/A)
Applied
100%
Developmental
(N/A)
Goals / Objectives
Salmonella enterica serotype Dublin is host-adapted to cattle and an important zoonotic risk. Most animals infected with S. Dublin have symptoms different than classical salmonellosis (respiratory symptoms instead of classic gastroenteritis), leading to substantial underdiagnosis. Herds affected by this pathogen face substantial costs associated with calf losses and are at a greater risk of being challenged by antimicrobial resistant pathogens. Most S. Dublin isolated to date have been reported to be multidrug resistant and to carry a mobile genetic element that can be passed to other bacteria.This strain was formerly believed to only exist in the western United States; however, a progressive increase in the isolation of S. Dublin from diagnostic samples in the eastern US has happened in recent years. However, no data are available regarding the herd prevalence of S. Dublin in Pennsylvania dairy herds, despite this pathogen being isolated from several human cases in Pennsylvania since 2014 and many animal cases in the past two years.The goal of this proposal is to address the potentially underdiagnosed problems caused by Salmonella Dublin in the Pennsylvania dairy industry by completing the following objectives:[1] Quantify the prevalence of Salmonella Dublin in Pennsylvania[2] Apply advanced detection methods to environmental samples[3] Develop mitigation efforts tailored to farms experiencing challenges by S. Dublin[4] Create training and extension materials about Salmonella Dublin-caused disease and prevention strategies. Materials will be widely disseminated through Penn State extension websites and presential events organized by the research team in collaboration with Penn State Extension.
Project Methods
Objective 1 Survey of Pennsylvania Dairy FarmsWe will perform ELISA antibody tests in samples from PA dairy farms to define baseline levels of seroprevalence for Salmonella Dublin. We have designed a multi-pronged recruitment and testing strategy to achieve this goal, given the varying herd sensitivity of different testing.Point Prevalence Estimation and Surveys: Raw milk and a corresponding management survey will be collected from each farm once to capture point prevalence of Salmonella Dublin in Pennsylvania. A combination test of a bulk tank milk ELISA and serology of animals with current or previous signs of Salmonellosis and/or youngstock ages 4-6 months will be carried out in select farms. Raw milk and blood samples will be tested with an indirect ELISA assay (PrioCHECK Salmonella Ab bovine Dublin milk cattle). Management surveys will be used to collect information on farm factors that are known to be risk factors for S. Dublin infection (e.g., herd size, off-site heifer rearing, purchase of animals, and calf management).Economic Analysis: Gross margin losses will be estimated by simulation following models published by Nielsen et al., 2013 and adapted to fit local dairy farm production settings.Analysis and Interpretation: Prevalence estimates will be adjusted for the percent of herds in each category as appropriate. Survey results will be summarized in tabular format and reported as means and standard deviations. Gross margin estimates will be calculated by using the model fitted in R, and different scenarios will be simulated by changing variables that are impacted by Salmonella Dublin infection (e.g. culling rate, medication costs). Estimates between different scenarios will be compared to calculate gross margin losses in each instance.Projected Outcomes: Herd-level prevalence of Salmonella Dublin among Pennsylvania dairy farms based on antibody testing of bulk tank milk and/or combination strategies. Management practices and their association with prevalence of Salmonella Dublin. Cost estimate of Salmonella Dublin introduction into Pennsylvania Dairy Farms. Increased awareness of the issue among extension personnel and dairy producers.Objective 2 Environmental Sampling Targeted for Detection of Salmonella DublinWe will test an improved protocol (Goodman et al., 2017) for detection and isolation of Salmonella Dublin in a select set of farms by using different approaches to test milk filters, manure, and calving area swabs, and milkhouse swabs in each farm in two distinct cross-sectional timepoints.Traditional Testing: Selective enrichment protocols carried out by the Animal Diagnostic Laboratory will be performed for bacterial isolation. Briefly, samples will be collected aseptically, placed in a sterile Whirl-Pak bag with evaporated skim milk and kept on ice for transportation to the laboratory. Testing will include Tetrathionate (TT) and ?Rappaport-Vassiliadis soya (RVS) broths incubated at 42±2° C for 20-24 hours. Confirmatory subcultures will be performed by subculturing the incubated RVS and TT broths onto brilliant green (BG), and/or XLT4 (Neogen) agar plates and incubating at 37±2°C for 20-24 hours. Colonies showing appropriate characteristics will be confirmed as belonging to the Salmonella genus via MALDI-TOF. Serogrouping will be performed by slide agglutination using commercial antisera. Antibiotic susceptibility testing will be performed using a Sensititre susceptibility system available at the Animal Diagnostic Laboratory.Enrichment qPCR: The protocol published from Goodman et al. 2017 will be followed for screening by qPCR. Briefly, total nucleic acid will be extracted from 1mL of RVS each enrichment and tested with ?Salmonella spp. detection kit (MicroSEQ Salmonella spp. Detection Kit, ThermoFisher Scientific). ?Internal positive amplification controls are included as part of the lyophilized reaction mix (run by multiplex). A negative amplification control (provided with the kit in a separate tube) will be included for each run.Salmonella CRISPRSeroSeq: To test the hypothesis that Salmonella Dublin is present in most samples but is often outcompeted by other flora during enrichments, we will select a subset of 20 enrichments confirmed positive for Dublin and 20 negative enrichments to perform Salmonella CRISPRSeroSeq in collaboration with Dr. Nikki Shariat at University of Georgia College of Veterinary Medicine.Analysis and Interpretation: Prevalence in herds identified as high-risk for Salmonella Dublin will be calculated as described in Objective 1. Agreement between the three methods will be calculated based on Cohen's kappa coefficients. Limit of agreement plots (Bland-Altman plots) will be calculated to determine if there are systematic differences between two sets of observations.Projected Outcomes: Prevalence of Salmonella Dublin in environmental samples in 5 dairy farms identified as being at high-risk of having S. Dublin, as measured with three different approaches. Advanced detection methods to help tailor mitigation strategies for a "seek and destroy" approach.Objective 3 Salmonella Dublin Spread MitigationWe will follow a structured approach based on risk scoring, testing, and management procedures developed by Danish epidemiologists as part of a Salmonella Dublin control program.Tailored mitigation plans: This approach relies both on risk assessment and targeted testing. This tool addresses the main points of concern regarding transmission of Salmonella Dublin (e.g., calving area, pre- and post- wean calf management, heifer rearing, cow health, and introduction of animals from other herds). An overall risk score is calculated based on responses to each area and provides an automated radar chart that highlights points of largest importance for mitigation efforts. Based on the risk identified, target bulk tank, environmental and/or animal sampling will be performed for detection of Salmonella Dublin. A management plan will be drafted for each farm with input from farm managers, and two consecutive visits will be performed 3 and 6 months after the initial visit to assess potential improvement on management practices and to collect subsequent samples for Salmonella Dublin detection.Projected Outcomes: Improved management and mitigation of Salmonella Dublin spread in positive herds. A tailored approach designed based on risk assessment in each herd. Improved management and biosecurity in S. Dublin positive herds in PA.Objective 4 Educate Dairy Veterinarians and Farmers About the Salmonella Dublin-caused Disease and Prevention StrategiesWe will leverage the extensive reach of Penn State Extension to provide educational resources for PA dairy farm owners and employees, as well as the veterinarians who work with these dairy farms. These resources and engagement opportunities will increase knowledge, and positively impact attitudes and behaviors about preventing and controlling S. Dublin in dairy herds and reducing the risk of zoonotic disease. Since veterinarians are highly trusted advisors and sources of information for dairy and livestock producers, they will be one of the audiences targeted, in addition to dairy farm owners and employees. A suite of extension resources will be developed to increase the level of knowledge about Salmonella Dublin, and to change attitudes and behaviors towards the prevention, detection, and control of Salmonella in dairy herds. These resources will include "Learn Now" videos, webinars, and on-farm workshops for herd owners and employees. These resources will be produced in English, with Spanish versions being made available to farm employees. In addition to our findings, the extension materials will include information concerning health risk, infection presentation, potential reservoirs, and risk management strategies.