Development of challenge models to evaluate the efficacy of a vaccine to reduce carriage of Salmonella in peripheral lymph nodes of cattle.

Because challenge models to infect peripheral lymph nodes (PLNs) with Salmonella have not been reported, we performed a series of experiments to develop and refine challenge models to evaluate an intervention applied at the animal level and to provide initial estimates of efficacy of an intervention (i.e., a vaccine) to aid in the design of future studies. In each of four experiments, steers (control or vaccinated) were inoculated with Salmonella strains Montevideo or Newport, and in experiment IV, Salmonella Senftenberg was also used. Calves were euthanized 14 to 42 days postinoculation, and PLNs were collected. In the first experiment, calves were challenged with ∼10¹° Salmonella cells, and few treatment differences were observed 14 days postchallenge. However, by day 21, Salmonella Newport was recovered from fewer vaccinated calves than control calves (P < 0.05). In experiment II, calves were challenged with ∼107 Salmonella cells and, after two necropsies (14 and 28 days postchallenge), only one lymph node was Salmonella positive; therefore, the study was terminated. In experiment III, calves were again challenged with ∼10¹° Salmonella cells, and no significant effect of vaccine was observed in calves challenged with Montevideo or Newport strains. A transdermal route of challenge was explored in experiment IV, using a 10-lancet, allergy testing instrument. Sixteen steers were challenged with either Salmonella Newport or Salmonella Montevideo (Salmonella Newport right legs; Salmonella Montevideo left legs), and all steers were challenged on the lower abdomen with Salmonella Senftenberg. Transdermal inoculation resulted in predictably Salmonella-positive PLNs, and a modest vaccine effect was detected. Because it is well tolerated by the calves and results in predictable and regionally specific Salmonella recovery from PLNs, the transdermal route of challenge may be preferred by researchers wishing to evaluate the impact of interventions designed to reduce the carriage of Salmonella in PLNs.

Evaluation of the potential antimicrobial resistance transfer from a multi-drug resistant Escherichia coli to Salmonella in dairy calves.

Previous research conducted in our laboratory found a significant prevalence of multi-drug resistant (MDR) Salmonella and MDR Escherichia coli (MDR EC) in dairy calves and suggests that the MDR EC population may be an important reservoir for resistance elements that could potentially transfer to Salmonella. Therefore, the objective of the current research was to determine if resistance transfers from MDR EC to susceptible strains of inoculated Salmonella. The experiment utilized Holstein calves (approximately 3 weeks old) naturally colonized with MDR EC and fecal culture negative for Salmonella. Fecal samples were collected for culture of Salmonella and MDR EC throughout the experiment following experimental inoculation with the susceptible Salmonella strains. Results initially suggested that resistance did transfer from the MDR E. coli to the inoculated strains of Salmonella, with these stains demonstrating resistance to multiple antibiotics following in vivo exposure to MDR EC. However, serogrouping and serotyping results from a portion of the Salmonella isolates recovered from the calves post-challenge, identified two new strains of Salmonella; therefore transfer of resistance was not demonstrated under these experimental conditions.

The effect of chlortetracycline on faecal microbial populations in growing swine.

The effect of antimicrobial use on the gastrointestinal microbiota of food animals is of increasing concern as bacteria accumulate resistance to multiple antimicrobials. Only a small fraction of the gastrointestinal microbiome is culturable, complicating characterisation of the swine gastrointestinal ecosystem. The objective of this study was to determine the effect of a growth promotion dose (50g/ton) of chlortetracycline on the phylogenetic diversity of bacteria from swine faeces using a culture-independent method. Four freshly weaned pigs were provided a grower ration of primarily corn (63.7%) and soybean meal (25.2%) for 21 days; on Day 21 for 4 weeks the diet of two pigs was medicated with 50g/ton chlortetracycline. Faecal material was collected from each pig on Days 0, 14, 23, 28, 35, 42 and 49 for 454-pyrosequencing of the 16S rRNA gene. UniFrac analysis of pyrosequencing data showed no significant difference in bacterial diversity based on diet and among pigs (P>0.05) fed the low-level dose of chlortetracycline. The most abundant phyla in both treatment groups were Firmicutes, Bacteroidetes, Proteobacteria and Spirochaetes. Higher concentrations of chlortetracycline (e.g. 200g/ton or 400g/ton) may be required to observe a shift in the gastrointestinal flora in swine faeces compared with the low-level dose in this study. Studies of broader scope are needed to understand thoroughly how growth-promoting antimicrobials influence the gut microflora and benefit food animal growth efficiency.

Lack of effect of feeding citrus by-products in reducing salmonella in experimentally infected weanling pigs.

The objective of the current research was to determine if feeding the citrus by-products(D) -limonene (DL) and citrus molasses would reduce the concentration and prevalence of Salmonella in weanling pigs experimentally infected with Salmonella Typhimurium. Twenty crossbred weanling pigs (average body weight [BW], 19.9 kg) were randomly assigned to one of four treatments: control, low-dose DL (1.5 ml/kg of BW per day), high-dose DL (3.0 ml/kg of BW per day), and citrus molasses (0.05 kg/kg of BW per day). Treatments were administered in the feed (twice daily) for 7 days, with one-half of the dose administered at each feeding. Fecal samples were collected twice daily (prior to administration of treatment) and cultured for quantitative and qualitative determination of the challenge strain of Salmonella. Upon termination of the study, pigs were euthanized and tissues from the stomach, ileum, cecum, spiral colon, and rectum, as well as luminal contents, were collected. In addition, the popliteal and ileocecal lymph nodes and liver, spleen, and tonsil tissue were collected for qualitative Salmonella culture. No significant treatment differences (P > 0.05) were observed among treatments for fecal concentration or prevalence of Salmonella throughout the 7-day collection period. Likewise, no treatment differences (P > 0.05) were observed for any of the tissue or luminal content samples collected. Salmonella was not cultured from the muscle-bound popliteal lymph node but was cultured from the mesenteric ileocecal lymph nodes. While there were no effects in the current experiment, future research may examine the effect of a lower challenge dose and/or different administration (dose or duration) of the citrus by-products.

Evaluation of the effect of gallium maltolate on fecal Salmonella shedding in cattle.

Strategies aimed at reducing fecal shedding of Salmonella and other foodborne pathogens may be effective for limiting transmission of pathogens from food animals to humans. The objective of this study was to determine the effectiveness of gallium maltolate (GaM) against Salmonella in vitro and to determine whether oral administration of GaM would reduce fecal shedding of Salmonella in cattle. Gallium is a semimetal exhibiting antimicrobial properties against some pathogenic bacteria, including Salmonella, by exploiting their need for iron to survive and replicate. In vitro growth studies were performed in pure cultures of Salmonella and in mixed cultures from ruminal fluid. Inclusion of GaM in culture medium or in mixed cultures of ruminal fluid resulted in a significant reduction in growth of Salmonella, suggesting that GaM may be effective for limiting growth and survival in vivo. Therefore, we subsequently administered two doses of GaM to Holstein steers, experimentally infected them with Salmonella, and quantitatively and qualitatively monitored fecal shedding at 12-h intervals. Sixty hours after beginning treatment, cattle were euthanized, and luminal contents and tissue were aseptically harvested from the rumen, jejunum, spiral colon, cecum, and rectum. The luminal contents were processed for quantitative and qualitative analysis of the challenge strains of Salmonella, and tissue samples were enriched and plated for qualitative analysis. We found no significant differences between control and treated animals in quantitative levels of Salmonella in the feces or the luminal contents. Likewise, we observed no pattern between control and treated animals in the frequency of positive or negative results from enriched feces, luminal contents, or tissue samples. These results suggest that GaM was not effective for reducing Salmonella in cattle.

Prediction of beef carcass salable yield and trimmable fat using bioelectrical impedance analysis.

Bioelectrical impedance technology (BIA) is capable of providing an objective method of beef carcass yield estimation with the rapidity of yield grading. Electrical resistance (Rs), reactance (Xc), impedance (I), hot carcass weight (HCW), fat thickness between the 12th and 13th ribs (FT), estimated percentage kidney, pelvic, and heart fat (KPH%), longissimus muscle area (LMA), length between electrodes (LGE) as well as three derived carcass values that included electrical volume (EVOL), reactive density (XcD), and resistive density (RsD) were determined for the carcasses of 41 commercially fed cattle. Carcasses were subsequently fabricated into salable beef products reflective of industry standards. Equations were developed to predict percentage salable carcass yield (SY%) and percentage trimmable fat (FT%). Resulting equations accounted for 81% and 84% of variation in SY% and FT%, respectively. These results indicate that BIA technology is an accurate predictor of beef carcass composition.

Acyl-homoserine-lactone autoinducer in the gastrointestinal [corrected] tract of feedlot cattle and correlation to season, E. coli O157:H7 prevalence, and diet.

Acyl-homoserine-lactone autoinducer (AHL) produced by nonenterohemorrhagic Escherichia coli species in cattle appears to be required for enterohemorrhagic E. coli (EHEC) colonization of the gastrointestinal tract (GIT). The current research aimed to examine the effect of season, diet, EHEC shedding, and location within the GIT on AHL prevalence in the ruminant. Luminal content samples were collected from the rumen and rectum of feedlot cattle at slaughter in the spring, summer, fall, and winter for culture of E. coli O157:H7 and AHL determination. During the spring collection, samples were additionally collected from the cecum and small intestine, but these samples all were AHL negative and therefore not examined again. To assess the influence of diet on AHL prevalence, 14 lambs were fed either 100% forage or 80% concentrate diets and experimentally inoculated with EHEC. At 8 days after infection, all the lambs were killed, and necropsies were taken, with luminal contents collected from the GIT. The collections from the feedlot cattle had AHL in 100% of the rumen content samples from the spring, summer, and fall, but not in any of the winter samples. No other GIT samples from feedlot cattle were AHL positive, and all the samples from the sheep study were AHL negative. The cattle seemed to show a weak correlation between ruminal AHL and EHEC prevalence. This research found AHL only in the rumen and not in the lower GIT of feedlot cattle. However, it is unclear whether this is because the pH of the lower gut destroys the AHL or because a lack of certain bacteria in the lower gut producing AHL.

An exploratory observational study to develop an improved method for quantifying beef carcass salable meat yield.

Eighty-seven grain-finished steers were harvested, evaluated, and fabricated into wholesale cuts to determine what measured composition indicators most accurately describe the percentage of closely trimmed salable meat yield. Indicators of lean and fat composition present at the cross-section between the 12th and 13th ribs were objectively evaluated using Assess image analysis software. Salable meat yield ranged from 50.18% to 72.92%, trimmable fat yield ranged from 12.87% to 36.69%, and bone yield ranged from 10.07% to 19.21%. Regression models were developed to estimate percentage of total salable meat yield. Composition indicators chosen to predict salable meat yield included hot carcass weight (HCW), perinephric fat weight, longissimus muscle area (LMA), subcutaneous fat thickness (SFT), ratio of LMA to subcutaneous fat area, and ratio of subcutaneous fat depth to HCW. These results indicate that prediction of beef carcass salable meat yield can be improved via modification to current measures used in the USDA yield grade equation and addition of new measures.

Technical note: the United States Department of Agriculture beef yield grade equation requires modification to reflect the current longissimus muscle area to hot carcass weight relationship.

With the adoption of visual instrument grading, the calculated yield grade can be used for payment to cattle producers selling on grid pricing systems. The USDA beef carcass grading standards include a relationship between required LM area (LMA) and HCW that is an important component of the final yield grade. As noted on a USDA yield grade LMA grid, a 272-kg (600-lb) carcass requires a 71-cm(2) (11.0-in.(2)) LMA and a 454-kg (1,000-lb) carcass requires a 102-cm(2) (15.8-in.(2)) LMA. This is a linear relationship, where required LMA = 0.171(HCW) + 24.526. If a beef carcass has a larger LMA than required, the calculated yield grade is lowered, whereas a smaller LMA than required increases the calculated yield grade. The objective of this investigation was to evaluate the LMA to HCW relationship against data on 434,381 beef carcasses in the West Texas A&M University (WTAMU) Beef Carcass Research Center database. In contrast to the USDA relationship, our data indicate a quadratic relationship [WTAMU LMA = 33.585 + 0.17729(HCW) -0.0000863(HCW(2))] between LMA and HCW whereby, on average, a 272-kg carcass has a 75-cm(2) (11.6-in.(2)) LMA and a 454-kg carcass has a 96-cm(2) (14.9-in.(2)) LMA, indicating a different slope and different intercept than those in the USDA grading standards. These data indicate that the USDA calculated yield grade equation favors carcasses lighter than 363 kg (800 lb) for having above average muscling and penalizes carcasses heavier than 363 kg (800 lb) for having below average muscling. If carcass weights continue to increase, we are likely to observe greater proportions of yield grade 4 and 5 carcasses because of the measurement bias that currently exists in the USDA yield grade equation.

Comparison of Gas Temperatures Measured by Coherent Anti-Stokes Raman Spectroscopy (CARS) of O(2) and N(2).

We investigate the accuracy of temperature measurements by coherent anti-Stokes Raman spectroscopy (CARS) of O(2) and use measurements taken with N(2) CARS and a thermocouple for comparison. Scanning vibrational CARS spectra of O(2) and N(2) were recorded over a broad range of temperatures: between 294 K and 1900 K in air that was heated in a tube furnace and at approximately 2450 K in a fuel-lean CH(4)-O(2)-N(2) flame. Temperatures were derived from least-squares fits of simulated and experimental spectra. Both the fundamental vibrational band and the first hot vibrational band were included in fitting. In the case of the tube furnace, the N(2) and the O(2) CARS temperature measurements agreed to within 3%, and results were similar with the thermocouple; in the flame the agreement was to within 1%. We conclude that, for cases in which O(2) is present in sufficient concentrations ( approximately 10% or greater), the accuracy of O(2) thermometry is comparable with that of N(2).

Dual-pump coherent anti-Stokes Raman scattering measurements of nitrogen and oxygen in a laminar jet diffusion flame.

Dual-pump coherent anti-Stokes Raman scattering (CARS) has been demonstrated for the simultaneous measurement of gas-phase temperature and concentrations of molecular nitrogen and oxygen. A polarization technique was used to vary the relative intensities of the two CARS signals and expand the dynamic range of the relative concentration measurements. Detailed temperature and oxygen mole fraction measurements were performed in the stabilization region of a hydrogen-nitrogen jet diffusion flame. These results indicate that there is a region below the nozzle exit where significant amounts of oxygen are found on the fuel side of the peak flame temperature profile.

Detection of methyl radicals in a flat flame by degenerate four-wave mixing.

We report the spatially resolved detection of methyl radicals in a methane-air flat flame, using degenerate four-wave mixing (DFWM). A frequency-tripled dye laser pumped with a frequency-doubled Nd:YAG laser was used to access the Herzberg beta(1) band of methyl near 216 nm. Using a nearly phase-conjugate geometry, we detected methyl with high spatial resolution [0.2 mm (0.3 mm) vertical (horizontal) and ~6 mm longitudinal] and with good signal-to-noise ratio in a rich (ø = 1.55) flame. Compared with laser absorption spectra, DFWM spectra were much less influenced by a broad featureless background. From the absorption data, we measured the peak methyl concentration to be 650 parts in 10(6), resulting in an estimated DFWM detection limit of 65 parts in 10(6).

Detection of trace molecular species using degenerate four-wave mixing.

Spectroscopies that make use of laser light have provided an important tool to modern researchers for the nonintrusive analysis of chemical systems. The strengths and limitations of these spectroscopic techniques often determine the viability of scientific investigations. The unique properties of degenerate four-wave mixing, a nonlinear optical technique, have recently been found to provide powerful capabilities for a wide range of applications.

Two-dimensional imaging of OH in flames by degenerate four-wave mixing.

Degenerate four-wave mixing is used to produce two-dimensional images of OH distributions in atmosphericpressure flames. The phase-conjugated images from single laser pulses exhibit excellent signal-to-noise ratios and illustrate that degenerate four-wave mixing has outstanding potential as a multidimensional diagnostic for combustion environments.

Nonlinear Gaussian expansions of stochastic processes: super-Gaussian effects in coherent anti-Stokes Raman spectroscopy.

We propose a new approach to the analysis of stochastic processes (such as fluctuating laser fields) with super-Gaussian statistics: the expansion of stochastic processes in terms of first and higher powers of Gaussian components, which are used like a basis set. This approach is applied to treat the super-Gaussian correlation effects observed in coherent anti-Stokes Raman scattering experiments using frequency-doubled pump laser fields. Our results give much better agreement with experimental data than previous theories do.

Comparison of motionally narrowed coherent anti-Stokes Raman spectroscopy line shapes of H(2) with hard- and soft-collision models.

We have compared high-resolution coherent anti-Stokes Raman spectroscopy measurements of the Q(1) transition of H(2) with theoretical line shapes incorporating both Doppler broadening and motional narrowing. Least-squares fits varying the motional narrowing coefficient for both the hard- and soft-collision model yielded line shapes that agreed well with observed spectra at 295 K for pressures between 50 and 3050 Torr. However, the narrowing coefficient for the hard-collision model was pressure dependent, whereas the narrowing coefficient for the soft-collision model was independent of pressure and yielded an optical diffusion coefficient that agreed with previously measured values.

High-resolution CARS measurements of temperature profiles and pressure in a tungsten lamp.

We use high-resolution coherent anti-Stokes Raman spectroscopy to measure the temperature profile and pressure in a nitrogen-filled tungsten lamp. Using a modified exponential-gap model for the J-dependent collisional-broadening coefficients, we obtain good fits to the experimental spectra, yielding spatially resolved temperatures and pressures with an accuracy of +/-5%.

High-resolution measurements of saturated coherent anti-Stokes Raman spectroscopy line shapes.

We report the first measurements to our knowledge of the effects of two-photon Raman saturation on fully resolved, homogeneously broadened coherent anti-Stokes Raman spectroscopy (CARS) line shapes. Saturation dips and line broadening were observed with a high-resolution scanning CARS experiment using a single-mode, frequency-doubled Nd:YAG laser and a pulse-amplified cw dye laser. Saturated line shapes are compared with the results of numerical solutions of the time-dependent density matrix equations.

Calculation of collisionally narrowed coherent anti-Stokes Raman spectroscopy spectra.

High-resolution coherent anti-Stokes Raman spectroscopy spectra of the N(2) Q branch at 294 K have been obtained at 1, 5, and 10 atm. Even at 1-atm pressure, disagreements with spectra calculated using the isolated line approximation were observed, indicating the importance of collisional narrowing effects in describing these spectra. A method of using the full G-matrix approach for the calculation of these spectra that is both exact and computationally efficient (requiring only one matrix diagonalization and inversion per spectrum) is discussed. Excellent agreement with experimental data is obtained using this method and a simple exponential gap model for the off-diagonal G-matrix elements.