Abstract

Bacterial attachment on surfaces is an important biological and industrial concern. Many parameters affect cell attachment behavior, including surface roughness and other topographical features. An understanding of these relationships is critical in the light of recent outbreaks caused by foodborne bacteria. Postharvest packing lines have been identified as a potential source of cross-contamination with pathogens, which can cause subsequent foodborne illness. The objective of this article is to evaluate the influence of surface topographical features on bacterial attachment at various processing temperatures to determine the extent of bacterial colonization. Type 304 stainless steel surfaces and pathogenic Listeria monocytogenes Scott A were used for a detailed investigation. Two commonly used surface types, extruded and ground, were evaluated to determine differences in bacterial attachment on the same type of material. Fifteen surface topography parameters at three processing temperatures were studied to evaluate possible correlations with microbial attachment on these surfaces. Scanning electron microscopy, energy-dispersive X-ray spectroscopy, and confocal microscopy were used for both qualitative and quantitative analyses of surfaces. An analysis of variance and multivariate regression analysis were used to predict the attachment behavior of L. monocytogenes Scott A on stainless steel surfaces. Surface isotropy, average surface roughness, surface spacing, and processing temperatures were strongly correlated with bacterial attachment on 304 stainless steel material.

Highlights

California has one of the largest systems of surface water reservoirs in the world, providing irrigation water to California's agriculturally productive Central Valley. Irrigation water is recognized as a vehicle for the microbial contamination of raw produce and must be monitored according to new federal regulation. The purpose of this study was to further understanding of the variability of fecal indicator bacteria (Escherichia coli and fecal coliforms) and pathogens (E. coli O157:H7 (O157), non-O157 Shiga toxin-producing E. coli (STEC) and Salmonella) along both horizontal and vertical profiles within California reservoirs. Monthly sampling was conducted in six reservoirs located in the foothills of the Western Sierra Nevada during the summer irrigation season and extreme drought conditions of 2014 (n = 257). Concentrations of fecal indicator bacteria were highly variable between reservoirs (p < 0.05) and along the horizontal profile (p < 0.001) from upstream to downstream, with higher concentrations typically found outside of the reservoirs than within. Though many of the reservoirs were thermally stratified, bacterial concentrations were not associated with water temperature (p > 0.05) or any one particular depth strata (p < 0.05). However, prevalence of Salmonella and STEC (16/70 and 9/70 respectively) was higher in the deep strata than in mid or surface layers. We found no statistical association between samples collected downstream of reservoirs and those from the reservoirs themselves. Continued monitoring and modeling of both bacterial indicators and enteric pathogens are critical to our ability to estimate the risk of surface irrigation water supplies and make appropriate management decisions.

In 2011, the US Congress passed the Food Safety Modernization Act, which tasks the US Food and Drug Administration to establish microbiological standards for agricultural water. However, little data are available for the microbiological quality of surface water irrigation supplies. During the 2015 irrigation season, we conducted a baseline study on the microbial water quality of large irrigation districts in California ( = 2) and Washington ( 4). Monthly samples ( 517) were analyzed for bacterial indicators (fecal coliforms, enterococci, and ) and pathogens ( spp., O157, and non-O157 Shiga toxin-producing [STEC]). Although there was a high degree of variability (μ ± SD = 59.13 ± 106.0), only 11% of samples (56/517) exceeded 126 colony-forming units (CFU) 100 mL, and only six samples exceeded 410 CFU 100 mL. Two volumes of water were collected for pathogen analysis (1 L and 10 L); prevalence of in 10-L samples (68149) was nearly double of that found in 1-L samples (132/517). We found STEC during ∼9% of sampling events (58/517); serotypes O26 and O45 were the most common at 31 and 26%, respectively. Pathogens were not associated with exceedance of the regulatory threshold, yet the odds of detecting increased approximately threefold (odds ration [O.R.] = 3.14, 0.0001) for every log increase in turbidity. Microbiological outcomes were highly district-specific, suggesting drivers of water quality vary across spatiotemporal scales. The true risk of contamination of produce from irrigation water supplies remains unknown, along with the optimal monitoring strategy to improve food safety.

The Sacramento-San Joaquin Delta Estuary (Delta) is the confluence of two major watersheds draining the Western Sierra Nevada mountains into the Central Valley of California, ultimately terminating into San Francisco Bay. We sampled 88 sites once a month for two years (2006-2008) over 87 separate sampling events for a total of 1740 samples. Water samples were analyzed for fecal indicator bacteria (Escherichia coli, enterococci and fecal coliforms), and 53 other physiochemical, land use, and environmental characteristics. The purpose of the study was to create a baseline of microbial water quality in the Delta and to identify various factors (climatic, land use, tidal, etc.) that were associated with elevated concentrations of indicator bacteria. Fecal indicator bacteria generally had weak to modest relationships to environmental conditions; the strength and direction of which varied for each microbial indicator, drainage region, and across seasons. Measured and unmeasured, site-specific effects accounted for large portions of variance in model predictions (ρ=0.086 to 0.255), indicating that spatial autocorrelation was a major component of water quality outcomes. The effects of tidal cycling and lack of connectivity between waterways and surrounding landscapes likely contributed to the lack of association between local land uses and microbial outcomes, though weak associations may also be indicative of mismatched spatiotemporal scales. The complex nature of this system necessitates continued monitoring and regular updates to statistical models designed to predict microbial water quality.

Field trials were conducted in July-August and October 2012 to quantify the inactivation rate of Escherichia coli O157:H7 when mixed with fecal slurry and applied to romaine lettuce leaves. Lettuce was grown under commercial conditions in Salinas Valley, California. One-half milliliter of rabbit, chicken, or pig fecal slurry, containing an average of 4.05 × 10⁷ CFU E. coli O157:H7 (C₀), was inoculated onto the upper (adaxial) surface of a lower leaf on 288 heads of lettuce per trial immediately following a 2.5 h irrigation event. To estimate the bacterial inactivation rate as a function of time, fecal matrix, irrigation and seasonal climate effects, sets of lettuce heads (n = 28) were sampled each day over 10 days and the concentration of E. coli O157:H7 (C_t) determined. E. coli O157:H7 was detected on 100% of heads during the 10-day duration, with concentrations ranging from ≤340 MPN/head (∼5-log reduction) to >3.45 × 10¹² MPN/head (∼5-log growth). Relative to C₀, on day 10 (C_{t = 12}) we observed an overall 2.6-log and 3.2-log mean reduction of E. coli O157:H7 in July and October, respectively. However, we observed relative maximum concentrations due to bacterial growth on day 6 (maximum C_{t = 8}) apparently stimulated by foliar irrigation on day 5. From this maximum there was a mean 5.3-log and 5.1-log reduction by day 10 (C_{t = 12}) for the July and October trials, respectively. This study provides insight into the inactivation and growth kinetics of E. coli O157:H7 on romaine lettuce leaves under natural field conditions. This study provides evidence that harvesting within 24 h post irrigation has the potential to increase the concentration of E. coli O157:H7 contamination, if present on heads of romaine lettuce; foliar irrigation can temporarily stimulate substantial regrowth of E. coli O157:H7.

A field trial was conducted in July 2011 to quantify the inactivation rate of Escherichia coli O157:H7 when mixed with fecal slurry and applied to romaine lettuce leaves. Lettuce was grown under commercial conditions in Salinas Valley, CA. One-half milliliter of rabbit fecal slurry, containing 6.3 × 107 CFU of E. coli O157:H7, was inoculated onto the upper (adaxial) surface of a lower leaf on 240 heads of lettuce within 30 min after a 2.5-h irrigation event. Forty-eight romaine lettuce heads were collected per event at 2.5 h (day 0.1), 19.75 h (day 0.8), 43.25 h (day 1.8), 67.25 h (day 2.8), and 91.75 h (day 3.8) postinoculation and were analyzed for the concentration of E. coli O157:H7 (Ct). E. coli O157:H7 was detected on 100% of collected heads in concentrations ranging from 340 to 3.40 × 1010 most probable number (MPN) per head. Enumeration data indicate substantial growth of E. coli O157:H7 postinoculation (2.5 h), leading to elevated concentrations, 1 to 3 log above the starting inoculum concentration (Co). By the end of the 92-h trial, we observed a net 0.8-log mean reduction of E. coli O157:H7 compared with Co; however, after accounting for the substantial bacterial growth, there was an overall 2.3-log reduction by the final sampling event (92 h). On the basis of two different regression models that used either the raw data for Ct or log-transformed values of Ct/Co during the period 2.5 to 91.75 h postinoculation, there was an estimated 76 to 80% reduction per day in bacterial counts; however, more accurate predictions of MPN per head of lettuce were generated by using non-log-transformed values of Ct. This study provides insight into the survival of E. coli O157:H7 transferred via splash from a contaminated fecal source onto produce during irrigation. Moreover, these findings can help generate inactivation times following a potential contamination incident.

Agricultural recovery basins are an important conservation practice designed to provide temporary storage of sediment and water on farms before low-volume discharge. However, food safety concerns have been raised regarding redistribution of captured sediment and water to fields used for human food production. The purpose of this study was to examine the potential microbiological risk that recovery basins may contribute to nearby produce fields and to evaluate characteristics that may influence or mitigate those risks. Water and sediment samples were collected from participating farms in three states and evaluated for bacterial indicators and pathogens over several months. Overall, 45% ( = 48) of water samples and less than 15% ( = 13) of sediment samples were positive for spp. In water samples, the occurrence of was positively associated with the use of surface water as a source of irrigation compared with groundwater as well as log-scale increases in concentration. In sediment samples, was associated with basin location (region) and basin fill levels. Sediment exposed to drying during dewatering had lower concentrations of indicator and a lower proportion of positives than submerged sediment from the same pond. Surrounding landscape characteristics, including vegetative coverage, proximity to livestock operations, and evidence of wildlife, were not correlated with pathogen occurrence in either sediment or water samples, suggesting that although habitat surrounding ponds may be an attractant to wildlife, those features may not contribute to increased pathogen occurrence in agricultural recovery basins.

Consumption of raw produce commodities has been associated with foodborne outbreaks in the United States. In a recent Centers for Disease Control and Prevention report outlining the incidence of food-related outbreaks from 1998 to 2008, produce of all kinds were implicated in 46% of illnesses and 23% of deaths. Methods that quickly identify fecal contamination of foods, including produce, will allow prioritization of samples for testing during investigations and perhaps decrease the time required to identify specific brands or lots. We conducted a series of trials to characterize the sensitivity and specificity of scent detection dogs to accurately identify fecal contamination on raw agricultural commodities (romaine lettuce, spinach, cilantro, and roma tomatoes). Both indirect and direct methods of detection were evaluated. For the indirect detection method, two dogs were trained to detect contamination on gauze pads previously exposed to produce contaminated with feces. For the direct detection method, two dogs were trained to identify fecal contamination on fresh produce. The indirect method did not result in acceptable levels of sensitivity except for the highest levels of fecal contamination (25 g of feces). Each dog had more difficulty detecting fecal contamination on cilantro and spinach than on roma tomatoes. For the direct detection method, the dogs exhibited >75% sensitivity for detecting ≥0.25 g of feces on leafy greens (cilantro, romaine lettuce, and spinach) and roma tomatoes, with sensitivity declining as the amount of feces dropped below 0.025 g. We determined that use of a scent detection dog to screen samples for testing can increase the probability of detecting ≥0.025 g of fecal contamination by 500 to 3,000% when samples with fecal contamination are rare (≤1%).

The objective of the study was to assess the microbiological safety of popular recreational swimming sites in Central California. Water samples were collected from eleven monitoring sites across the lower reaches of two watersheds for two consecutive swimming seasons (2012-2013), and levels of indicator and pathogenic microorganisms were determined. Data on ambient weather and water chemistry were collected for analyzing their associations with microorganisms in water. All water samples were positive for indicator E. coli with mean concentrations per site ranging from 3.07 to 216.11 MPN/100 ml in 2012 and 13.4 to 226.97 MPN/100 ml in 2013. Mean E. coli concentrations in 27% and 36% samplings sites exceeded the EPA 2012 Recreational Water Quality Criteria recommended mean concentration of ≤ 126 CFU/100 ml of E. coli, in 2012 and 2013, respectively. Cryptosporidium spp. oocysts were detected in all water samples from all sampling sites, with an overall prevalence of 50% and mean concentrations of 0.08 oocysts/l in 2012 and 0.19 oocysts/l in 2013. Giardia spp. cysts were detected at eight sites, with an overall prevalence of 28.8% and mean concentration of 0.2 cysts/l in both years. The majority of the detected Cryptosporidium spp. oocysts and Giardia spp. cysts appeared damaged under microscopy. E. coli O157:H7 was detected in 9% of water samples, with positive samples limited to three sites. Salmonella spp. were detected in all but one site across the two years, with mean concentrations of 0.94 MPN/l in 2012 and 1.85 MPN/l in 2013. Cryptosporidium spp. oocyst concentrations were negatively associated with 30-day mean wind speed and cumulative precipitation and dissolved oxygen in water. Giardia spp. cyst concentrations were positively associated with turbidity and pH of water and negatively associated with E. coli concentrations and 24-h mean air temperature. Salmonella spp. concentrations were positively associated with 30-day mean air temperature. The occurrence of E. coli O157:H7 was positively associated with previous 30-day cumulative precipitation.