Prevalence of Campylobacter species and associated risk factors for contamination of dairy products collected in a dry season from major milk sheds in Ethiopia.

A cross-sectional study was conducted to investigate the prevalence and risk factors for contamination of Ethiopian dairy products with Campylobacter. A total of 912 dairy food samples were collected from establishments of 682 study participants that were interviewed. Samples were tested for Campylobacter by following the ISO 10272-1:2017 standard and PCR confirmation. Campylobacter was detected in 11% of tested food samples and all detected Campylobacter were C. jejuni. The highest prevalence of C. jejuni was found in raw milk (16%), followed by pasteurized milk (9%) and cottage cheese (2%) (P < 0.001). Using warm water and soap for cleaning cow udders and teats on farms reduced the likelihood of detecting Campylobacter in milk (AOR = 0.3, P = 0.023). Filtering milk with a cloth, using a plastic filter (AOR = 0.065, P = 0.005), and storing milk in an aluminum container (AOR = 0.23, P = 0.027) reduced the likelihood of detecting Campylobacter in milk at the collection facilities. In contrast, Campylobacter detection was significantly more likely in milk collected at collection centers with concrete floors (AOR = 5.2, P = 0.004). The odds of detecting Campylobacter in milk were 17 times greater (AOR = 17, P = 0.007) in milk processing facilities that did not calibrate a pasteurizer on an annual basis. Finally, having a separate refrigerator for milk storage reduced the odds of detecting Campylobacter in retail (AOR = 0.29, P = 0.021).

The use of a CDC biofilm reactor to grow multi-strain Listeria monocytogenes biofilm.

Listeria monocytogenes is one of the most concerning pathogens for the food industry due to its ability to form biofilms, particularly in difficult-to-clean sites of processing facilities. There is a current industry-wide lack of data to refer to when selecting a strategy to control L. monocytogenes biofilms in the food premises. Many strategies have been developed to study biofilm formation of bacteria; however, few have targeted L. monocytogenes biofilms under dynamic conditions. This study addresses the biofilm formation ability of L. monocytogenes on stainless steel and polycarbonate under dynamic conditions using TSBYE or BHI as media culture at 30 °C or 37 °C. Higher cell counts were recovered at 30 °C in TSBYE on polycarbonate while lower counts were obtained at 37 °C in BHI on stainless steel (P < 0.05). Nonetheless, all factors (temperature, media and material) were statistically significant (P < 0.05) and an interaction between temperature and media was observed (P < 0.05). To our knowledge, this work represents an initial framework to develop L. monocytogenes biofilms under different dynamic conditions. The use of CDC Biofilm Reactor is not widely used yet in the food industry and represent a novel approach to help sanitary control strategies implementation.

Review of the prevalence of foodborne pathogens in milk and dairy products in Ethiopia.

Food safety is a significant barrier to social and economic development throughout the world, particularly in developing countries. Here, we reviewed the prevalence of major bacterial foodborne pathogens (Salmonella spp., Listeria monocytogenes, Escherichia coli O157:H7 and Campylobacter spp.) in the rapidly growing Ethiopian dairy supply-chain. We identified 15, 9, 5 and 0 studies that had reported the prevalence of Salmonella spp., L. monocytogenes, E. coli O157:H7, and Campylobacter spp. in dairy foods, respectively. The studies reviewed reported a median prevalence of Salmonella, L. monocytogenes, and E. coli O157:H7 of 6, 9 and 10%, respectively, in raw cow milk in Ethiopia, indicating a concerning occurrence of bacterial foodborne pathogens in raw milk. Implementation of good hygiene and production practices and assessment of interventions targeting the reduction of contamination in the dairy supply chain is needed to inform coordinated efforts focused on improvement of dairy food safety in Ethiopia.

Seasonal variation of Salmonella enterica prevalence in milk and cottage cheese along the dairy value chain in three regions of Ethiopia.

Seasonal fluctuations influence foodborne illness transmission and affect patterns of microbial contamination of food. Previous investigations on the seasonality of Salmonella enterica prevalence in dairy products in Ethiopia have been minimal. However, such data are needed to inform strategic development of effective interventions to improve food safety, as seasonal differences may affect intervention strategies. This study was conducted to identify differences in the prevalence of Salmonella in milk and cheese samples between wet and dry seasons. A longitudinal study design was utilized with a random sampling occurring during both dry and wet seasons. A total of 448 milk and cottage cheese samples were collected from Oromia, Sidama, and Amhara regions. Samples were tested for Salmonella using the ISO 6579-1: 2008 method, followed by PCR confirmation. A chi-square test was conducted to assess the significance of differences in the prevalence of Salmonella in the samples between the two seasons. Results from this study showed a higher prevalence of Salmonella in all sample types during the dry season (P < 0.05). Moreover, when comparing raw milk, pasteurized milk, and cottage cheese samples, a significant difference was observed in Salmonella prevalence from raw milk samples (27.08%) collected in the Oromia region. Additionally, data showed a significantly higher prevalence of Salmonella in samples collected from raw milk producers (29.17%) during the wet season (P < 0.05). This study indicates that in order to enhance the safety of dairy products in Ethiopia, comprehensive, long-term awareness building on hygienic milk production and handling that consider seasonal influence is warranted. Supplementary Information: The online version contains supplementary material available at 10.1186/s40550-024-00108-4.

Evaluation of Methods for Identifying Poultry Wing Rinses With Salmonella Concentrations Greater Than or Equal to 10 CFU/mL.

In the United States, the Proposed Regulatory Framework to Reduce Salmonella Illnesses Attributable to Poultry published by the Food Safety and Inspection Service (FSIS) has highlighted the need for simple, rapid methods that identify poultry wing rinse samples harboring Salmonella concentrations ≥10 CFU/mL. One of eight cold-stressed and nutrient-starved Salmonella strains was inoculated into post-chill two-joint poultry wing rinses (48 turkey and 72 chicken) at levels from 0.22 to 3.79 log CFU/mL, and then measured by 3-tube Most Probable Number (MPN), BioMerieux GENE-UP QUANT, Hygiena BAX SalQuant, and novel threshold methods. The MPN lower limit of quantification (LLQ) for Salmonella was -0.96 log CFU/mL. MPN overestimated the inoculated Salmonella level by 0.05 ± 0.35 log CFU/mL. The GENE-UP QUANT Salmonella method (LLQ = 1.00 log CFU/mL) underestimated the inoculated Salmonella level by 0.05 ± 0.51 log CFU/mL. The BAX SalQuant method (LLQ = 0.00 log CFU/mL) underestimated the inoculated Salmonella level by 1.21 ± 0.78 log CFU/mL. Threshold test methods with Poisson probabilities of 0.95 (PiLOT-95), 0.86 (PiLOT-86), 0.63 (PiLOT-63), and 0.50 (PiLOT-50) were developed to identify poultry wing rinses harboring Salmonella levels ≥10 CFU. MPN was 93.1%, accurate for determining if Salmonella levels in poultry wing rinses were ≥10 CFU/mL, but MPN costs and time requirements can be prohibitive for most laboratories. GENE-UP quantification was 86.1% accurate, but the GENE-UP method requires equipment and technical expertise that some food safety laboratories may not possess. BAX quantification had the lowest accuracy; 58.4%. PiLOT threshold test accuracies ranged from 83.2% for PiLOT-50 to 93.1% for PiLOT-86. The PiLOT threshold tests are simple and can be adapted to identify many environmental or food samples containing Salmonella exceeding any user-defined concentration threshold.

Effect of accelerated aging on shelf-stability, product loss, sensory and biochemical characteristics in 2 lower quality beef cuts.

The aim of this study was to determine the impact of accelerated aging (AA) on shelf stability, product loss, sensory and biochemical characteristics of 2 lower quality beef cuts. Triceps brachii (TB) and semimembranosus (SM) were collected and fabricated from 10 USDA Choice beef carcasses and assigned to 1 of 6 treatments: 3 d cooler aged (control), 21 d cooler aged, AA 49 °C for 2 h, AA 49 °C for 3 h, AA 54 °C for 2 h, and AA 54 °C for 3 h. The results showed that AA can decrease APC counts on steak surface and in purge and redness, but increase lightness and product loss of the steaks (P < 0.01). Lower shear force was also found for AA steaks compared to those from the control (P < 0.01), with the AA 54 °C treatments being comparable to 21 d cooler aging. However, the trained sensory panel determined AA steaks were less juicy and flavorful than those from the control and 21 d cooler aged samples (P < 0.05). There was no off-flavor detected in AA steaks though lipid oxidation was higher in AA samples than those in the control steaks (P < 0.01). The AA treatments stimulated cathepsin activity (P < 0.05), which may have enhanced the solubilization of stromal proteins and led to a different troponin-T degradation pattern compared to those from the 21 d aged samples (P < 0.01). Although AA is an economical and time-efficient method to increase tenderness of lower-quality beef cuts, further research is needed to determine strategies to mitigate the decrease in juiciness from AA treatments.

Hygienic status of beef butcher shop facilities and antibiotic resistance profile of Salmonella enterica in Ethiopia.

The microbiological quality of meat is influenced by the conditions of hygiene prevailing during production and handling. Thus, this study aimed to assess the prevalence of Salmonella enterica and its antimicrobial resistance, load of hygiene indicator bacteria including E. coli (ECC), coliforms (CC), total coliform (TCC), Enterobacteriaceae (EB) and aerobic plate count (APC), and meat handler's food safety knowledge and hygiene practices in butcher shops in two cities, Addis Ababa and Hawassa in Ethiopia, during 2020 and 2021. A total of 360 samples of beef carcasses (n = 120), knives (n = 60), chopping boards (n = 60), weighing balance (n = 60), and personnel's hands (n = 60) were randomly collected for microbial analysis. Besides, 120 participants were selected to participate in a food safety knowledge and hygiene practices assessment. The S. enterica isolates were identified by agglutination test followed by qPCR targeting invA gene. Phenotypic antimicrobial resistance profiles of S. enterica were determined using disk diffusion assays as described in CLSI. The ECC, CC, TCC, EB, and APC populations were quantified by plating onto petrifilm plates. A structured questionnaire was used to determine food safety knowledge and hygiene practices of participants. Overall prevalence of S. enterica was 16.7% (95% CI, 8.3-26.7) and location seems to have no effect (p = 0.806). Only 20% of the S. enterica were resistant to ampicillin and tetracycline. However, the majority (80%) of S. enterica isolates were susceptible to the panel of 11 antimicrobials tested. The overall mean ± SD (log CFU/cm2) of ECC, CC, TCC, EB, and APC were 4.31 ± 1.15; 4.61 ± 1.33; 4.77 ± 1.32; 4.59 ± 1.38 and 5.87 ± 1.52, respectively. No significant difference (p = 0.123) in E. coli contamination was observed between samples of beef carcasses and chopping boards. The EB contamination showed no significant difference (p > 0.05) among sample sources. The APC contamination levels on beef carcass were significantly higher (p > 0.05) than other sample sources. A total of 56% (95% CI: 46.7 - 65.0) of the participants had poor knowledge and 65% (95% CI: 56.7 - 73.3) had poor hygiene practices towards food safety. This study highlighted the poor hygiene status of butcher facilities with a potential risk of beef safety. Thus, appropriate food safety control strategies and inspection is needed at retail establishments.

Genomic characterization of Listeria monocytogenes and Listeria innocua isolated from milk and dairy samples in Ethiopia.

Listeriosis caused by Listeria monocytogenes often poses a significant threat to vulnerable populations. Dairy products have been implicated in outbreaks of listeriosis worldwide. In Ethiopia, studies have identified Listeria spp. and L. monocytogenes in various dairy products, but the genetic diversity and phylogenetic relationships of these bacteria remain largely unknown in the low- and middle-income countries. Therefore, we conducted whole-genome sequencing on 15 L. monocytogenes and 55 L. innocua isolates obtained from different levels of the dairy supply chains across three regions in Ethiopia. Genomes were assembled and used for MLST genotyping and single nucleotide polymorphism (SNP) analysis to infer phylogenetic relationships. We identified a total of 3 L. monocytogenes (i.e., 2, 145, and 18) and 12 L. innocua (i.e., 1489, 1619, 603, 537, 1010, 3186, 492, 3007, 1087, 474, 1008, and 637) MLST sequence types among the studied isolates. Some of these sequence types showed region-specific occurrence, while others were broadly distributed across regions. Through high-quality SNP analysis, we found that among 13 L. monocytogenes identified as ST 2, 11 of them were highly similar with low genetic variation, differing by only 1 to 10 SNPs, suggesting potential selection in the dairy food supply chain. The L. innocua isolates also exhibited low intra-ST genetic variation with only 0-10 SNP differences, except for the ST 1619, which displayed a greater diversity.

Salmonella Prevalence and Quantification in Market Hog Lymph Nodes and Tonsils in Several Regions and Seasons of the United States.

Market hog lymph nodes (LNs) can contaminate carcasses with Salmonella, as well as ground and comminuted pork products. The objective of this study was to perform a qualitative and quantitative analysis of LNs from several regions and seasons in the United States to establish a Salmonella prevalence and concentration baseline. Six types of LNs (axillary, mesenteric, subiliac, tracheobronchial, superficial inguinal, prescapular), and tonsils were sampled from market hog carcasses from different regions (east, central, and west) and seasons (winter, spring, and summer/fall). Salmonella was detected and enumerated using BAX®-System-SalQuant® methods and the BAX®-System Real-Time Salmonella Assay. Salmonella prevalence (N = 4,132) was 36% for tonsils, 35% for mesenteric LN, and less than 10% for the other LN types. Of the 601 carcasses tested, 62% were positive for Salmonella, with the highest prevalence occurring during spring in the east (90.9%), and the lowest prevalence occurring during spring in the central region (26.0%). Tonsil prevalence was greatest in the eastern region during spring. Mesenteric LN prevalence was high (>20%) regardless of season or region. Salmonella prevalence in tracheobronchial, subiliac, axillary, and superficial inguinal LNs was generally greatest during the spring or fall and in the eastern region. The median SalQuant® Salmonella concentration was 2.18 log10Salmonella cells/sample. Median SalQuant® concentration for all other sample types fell below the limit of quantification (1 log10Salmonella cells/sample). This longitudinal study can be used by the pork industry for risk assessments and risk-based decision-making.

Seasonal Comparison of Microbial Hygiene Indicators in Raw and Pasteurized Milk and Cottage Cheese Collected across Dairy Value Chain in Three Regions of Ethiopia.

A longitudinal design with a simple random sampling method was used to collect and compare microbial hygiene levels between the dry season (January to April) and wet season (June to August). A total of 456 milk and cottage cheese samples were collected from each site along the dairy value chain from three regions. Enumeration of total aerobic mesophilic bacteria (APC), total coliforms (TCC), and Escherichia coli (EC) was performed according to standard methods. Independent t-tests were employed to assess the significant variation at (p < 0.05) between the two seasons. The cumulative result of APC of 7.61 log cfu/mL and g and TCC of 3.50 log cfu/mL in the dry season were significantly higher than the wet season of 7.15 log cfu/mL and 2.49 log cfu/mL, respectively, whereas generic E. coli count (EC) was significantly higher in the wet season (0.70 log cfu/mL and g) than that in the dry season (0.40 log cfu/mL and g). The results of hygienic indicator microbial load significantly varied with season. Hence, hygienic milk production and handling practices that comprehend seasonal influence should be implemented to improve the safety of milk.

Prevalence and associated risk factors for Salmonella enterica contamination of cow milk and cottage cheese in Ethiopia.

Animal sourced foods, such as dairy products, are common sources of Salmonella enterica, a foodborne pathogen of increasing global concern, particularly in developing countries. In Ethiopia, most data on the prevalence of Salmonella in dairy products is highly varied and limited to a specific region or district. Furthermore, there is no data available on the risk factors for Salmonella contamination of cow milk and cottage cheese in Ethiopia. This study was therefore conducted to determine the presence of Salmonella throughout the Ethiopian dairy value chain and to identify risk factors for contamination with Salmonella. The study was carried out in three regions of Ethiopia, including Oromia, Southern Nations, Nationalities and Peoples, and Amhara during a dry season. A total 912 samples were collected from milk producers, collectors, processors, and retailers. Samples were tested for Salmonella using the ISO 6579-1: 2008 method, followed by PCR confirmation. Concurrent with sample collection, a survey was administered to study participants to identify risk factors associated with Salmonella contamination. Salmonella contamination was highest in raw milk samples at the production (19.7%) and at milk collection (21.3%) levels. No significant difference in the prevalence of Salmonella contamination among regions was detected (p > 0.05). Regional differences were apparent for cottage cheese, with the highest prevalence being in Oromia (6.3%). Identified risk factors included the temperature of water used for cow udder washing, the practice of mixing milk lots, the type of milk container, use of refrigeration, and milk filtration. These identified factors can be leveraged to develop targeted intervention strategies aimed at reducing the prevalence of Salmonella in milk and cottage cheese in Ethiopia.

Genotypic and Phenotypic Characterization of Antimicrobial and Heavy Metal Tolerance in Salmonella enterica and Escherichia coli Isolates from Swine Feed Mills.

Antimicrobials and heavy metals are commonly used in the animal feed industry. The role of in-feed antimicrobials on the evolution and persistence of resistance in enteric bacteria is not well described. Whole-Genome Sequencing (WGS) is widely used for genetic characterizations of bacterial isolates, including antimicrobial resistance, heavy metal tolerance, virulence factors, and relatedness to other sequenced isolates. The goals of this study were to i) use WGS to characterize Salmonella enterica (n = 33) and Escherichia coli (n = 30) isolated from swine feed and feed mill environments; and ii) investigate their genotypic and phenotypic antimicrobial and heavy metal tolerance. Salmonella isolates belonged to 10 serovars, the most common being Cubana, Senftenberg, and Tennessee. E. coli isolates were grouped into 22 O groups. Phenotypic resistance to at least one antimicrobial was observed in 19 Salmonella (57.6%) and 17 E. coli (56.7%) isolates, whereas multidrug resistance (resistant to ≥3 antimicrobial classes) was observed in four Salmonella (12%) and two E. coli (7%) isolates. Antimicrobial resistance genes were identified in 17 Salmonella (51%) and 29 E. coli (97%), with 11 and 29 isolates possessing genes conferring resistance to multiple antimicrobial classes. Phenotypically, 53% Salmonella and 58% E. coli presented resistance to copper and arsenic. All isolates that possessed the copper resistance operon were resistant to the highest concentration tested (40 mM). Heavy metal tolerance genes to copper and silver were present in 26 Salmonella isolates. Our study showed a strong agreement between predicted and measured resistances when comparing genotypic and phenotypic data for antimicrobial resistance, with an overall concordance of 99% and 98.3% for Salmonella and E. coli, respectively.

A case report of Salmonella enterica serovar Corvallis from environmental isolates from Cambodia and clinical isolates in the UK.

Salmonella enterica subspecies enterica serovar Corvallis (S. Corvallis) has been identified as a human pathogen and as a food contaminant. Diarrhoeal disease is a common diagnosis in tourists visiting Southeast Asia, often with unknown aetiology. However, numerous public health institutes have identified Salmonella as a common causative agent when consuming contaminated food and water. Genomic data from environmental isolates from a Cambodian informal market were uploaded to the National Center for Biotechnology Information (NCBI) platform, allowing the novel sequences to be compared to global whole-genome sequence archives. The comparison revealed that two human clinical isolates from England and four of the environmental isolates were closely related, with an average single nucleotide polymorphism (SNP) difference of 1 (0-3 SNPs). A maximum-likelihood tree based on core SNPs was generated comparing the 4 isolates recovered from a Cambodian informal market with 239 isolates of S. Corvallis received from routine surveillance of human salmonellosis in England and confirmed the close relationship. In addition, the environmental isolates clustered into a broader phylogenetic group within the S. Corvallis population containing 68 additional human isolates, of which 42 were from patients who reported recent international travel, almost exclusively to Southeast Asia. The environmental isolates of S. Corvallis isolated from an informal market in Cambodia are concerning for public health due to their genetic similarity to isolates (e.g. clinical isolates from the UK) with known human virulence and pathogenicity. This study emphasizes the benefits of global and public data sharing of pathogen genomes.

Investigation on Salmonella enterica, Escherichia coli, and coliforms in beef from Ethiopian abattoirs: A potential risk of meat safety.

The aims of this study were to determine the prevalence of Salmonella enterica (S. enterica) antimicrobial resistance based upon phenotypic assessment, and level of sanitation indicator organisms from 150 beef carcasses collected from three representative abattoirs in eastern, central, and southern Ethiopia. Samples were screened for S. enterica prevalence following the U.S. Department of Agriculture Microbiology Laboratory Guidebook (MLG) and confirmed by real-time PCR. The S. enterica isolates were phenotypically evaluated for susceptibility to a panel of 13 antimicrobials using disk diffusion method as described in Clinical and Laboratory Standard Institutes guidelines. In addition, the coliform, total coliform, and generic E. coli populations were quantified by plating onto E. coli/coliform petrifilms. A total of 45 isolates of S. enterica were confirmed using real-time PCR. The overall prevalence of S. enterica at the carcass level was 22.7% (95% CI, 16.0-30.0). However, based on the sampling points, S. enterica prevalence detected on hides was 13.3% (95% CI, 8.0-18.7), pre-evisceration was 12.0% (95% CI, 7.3-17.3), and post-evisceration was 4.7% (95% CI, 1.3-8.0). The prevalence of S. enterica was significantly lower on post-evisceration as compared to hides (p = .009) and pre-evisceration (p = .022). No differences were detected for the prevalence of S. enterica between abattoirs (p = .346). Finally, the prevalence of S. enterica was highest during the wet season (p = .011). The overall mean log CFU/cm2 ± SD of generic E. coli, coliform, and total coliform counts were 4.55 ± 0.99, 4.91 ± 1.13, and 4.98 ± 1.09, respectively. About 20% of S. enterica exhibited phenotypic multidrug resistant. The most frequently detected resistance was to Tetracycline (28.9%), followed by Streptomycin (22.2%) and Sulfisoxazole (20.0%). The data from this study highlight the need to implement interventions on improvement of sanitary practices in abattoirs.

Effects of pigeon pea leaves and concentrate mixture on feed intake, milk yield, and composition of crossbred dairy cows fed native pasture hay.

Protein supplements are expensive and not easily accessible under small-scale livestock production systems in Ethiopia and other developing countries, which necessitates investigating the alternative protein sources for cost-effective livestock production. Pigeon pea (Cajanus cajan L. Millsp) leaves (PPLs) are rich in protein and are well-suited for feeding small ruminants; however, the effect of inclusion of PPL in the concentrate mixture (CM) on the performance of dairy cows was not well documented. This experiment was conducted to evaluate the effect of supplementation of PPL and CM to native pasture hay-based rations on feed intake, milk yield and composition, and blood metabolites of crossbred dairy cows (Holstein × Zebu). A 4 × 4 Latin square design with three replications, balanced for carryover effects, was used for this study. The treatments included native pasture hay provided ad libitum as a basal diet, supplemented with a CM alone (T1), the inclusion of 10% of PPL in the CM (T2), 20% PPL in the CM (T3), or 30% PPL in the CM (T4). Supplements were isocaloric and isonitrogenous. Total DM intake (hay + supplement intake) was similar (P > 0.05) among treatments. Hay intake was greater (P = 0.05) for T1 and T2 than for T4, while supplement intake was the least for T1 (P < 0.05). The treatment groups T2, T3, and T4, where PPL was included, had similar (P > 0.05) supplement intake. Feed intake, milk yield and composition, feed conversion efficiency, body condition score, serum total protein, albumin, globulin, glucose, triglyceride, urea N, creatinine, and cholesterol were similar (P > 0.05) among treatments. The inclusion of up to 30% of PPL in the CM resulted in a comparable performance of crossbred dairy cows as supplementation with CM under the conditions of the current experiment. Therefore, further study is required to evaluate the effect of the inclusion of a higher level of PPL in the concentrate mixture on the performance of lactating crossbred dairy cows.

Effects of moisture and temperature on Salmonella survivability in beef tallow, white grease, and chicken rendered fat.

High moisture levels introduced to fats after the rendering process can lead to Salmonella presence and growth. Limited research on strategies to eliminate pathogens in these environments are available. Rendered fat characteristics, such as water activity and fatty acids composition, may contribute to Salmonella survivability. The purpose of this research was to evaluate the effects of moisture levels (0%, 0.5%, 1%, and 3%), storage temperatures (48 and 76 ˚C), and fat characteristics on the growth and survival of Salmonella in beef tallow, white grease and poultry fat samples. Samples were inoculated with a high (~108 CFU/mL) and a low (~105 CFU/mL) Salmonella cocktail (S. Senftenberg, S. Newport, S. Thompson and S. Infantis). Samples were stored for up to 5 days at 48 and 76 ºC. Remaining population was evaluated daily with and without enrichment step. Death rates were calculated using Weibull model for each temperature and moisture level. Only temperature had an effect (P < 0.05) on Salmonella inactivation, while no effect between moisture and/or inoculum level were observed. When all products were challenged at 76 ˚C, counts were below detectable limits after 6 hours. At 48˚C a progressive decline in Salmonella population was observed within 3 days for both beef tallow and white grease when high inoculum was used for the challenge study. Salmonella was below detectable limit within 4 days for both fat types when a low inoculum was instead applied. This research identified the effect of moisture and temperature in rendered fat samples contaminated with Salmonella and underlined the need to use time-moisture-temperature data to minimize microbial growth during transportation and storage.

Prevalence of Salmonella enterica Isolated from Food Contact and Nonfood Contact Surfaces in Cambodian Informal Markets.

ABSTRACT: The lack of hygiene and sanitation practices and insufficient infrastructure in Cambodian informal markets may increase the risk of food contamination, specifically raw vegetables, which in turn may increase the chances of contracting a foodborne disease. The aims of this study in informal markets in Cambodia were (i) to quantify the prevalence of Salmonella enterica based upon differences in season of the year (rainy versus dry), surface types (food contact surfaces versus nonfood contact surfaces), and location of vendors within the market (inside versus outside) and (ii) to characterize S. enterica serotype prevalence. A total of 310 samples were screened for S. enterica prevalence following the U.S. Department of Agriculture guidelines, and results were confirmed by PCR assay. Whole genome sequencing was used to determine the serotype for each isolate in silico using SeqSero 1.0 on draft genomes. A total of 78 samples were confirmed positive for S. enterica. During the dry season, S. enterica was more prevalent on food contact surfaces than on nonfood contact surfaces (estimated probability of detection [confidence interval]: 0.41 [0.25, 0.59] and 0.17 [0.08, 0.32], respectively; P = 0.002), but no differences were apparent in the rainy season. No differences in S. enterica prevalence were found based on location within the market (P = 0.61). Sixteen S. enterica serotypes were detected across multiple surfaces. The most common S. enterica serotypes were Rissen (18 isolates), Hvittingfoss (11), Corvallis (10), Krefeld (8), Weltevreden (6), and Altona (6). Accurate data on the prevalence of S. enterica in informal markets are crucial for the development of effective surveillance and implementation of suitable intervention strategies at the domestic level, thus preventing foodborne illness.

Investigating Salmonella enterica, Escherichia coli, and Coliforms on Fresh Vegetables Sold in Informal Markets in Cambodia†.

ABSTRACT: Vegetables in Cambodia are commonly sold in informal markets lacking food safety standards and controls. Current data on microbial contamination of vegetables in Cambodian informal markets are limited. The purpose of this study was to investigate Salmonella enterica and indicator organisms (Escherichia coli and coliforms) on the surface of fresh vegetables sold in informal markets in Cambodia. Samples of loose-leaf lettuce, tomatoes, and cucumbers were collected from 104 vendors at four informal markets in Battambang and Siem Reap provinces during the rainy and dry seasons. Detection methods for S. enterica were adapted from the U.S. Food and Drug Administration's Bacteriological Analytical Manual. Coliform and E. coli populations were quantified by plating onto E. coli/coliform count plates. S. enterica was most prevalent on lettuce during the dry season (56.5%, 95% confidence interval [CI] [41.0, 70.8]) than during the rainy season (15.4%, 95% CI [7.5, 29.1]), whereas no significant seasonal differences were apparent for tomatoes and cucumbers. Regardless of season, levels of S. enterica were highest on lettuce (5.7 log CFU/g, 95% CI [5.5, 5.9]), relative to cucumbers (4.2 log CFU/g, 95% CI [3.8, 4.6]) and tomatoes (4.3 log CFU/g, 95% CI [4.1, 4.6]). For E. coli, prevalence was higher during the rainy season (34.0%, 95% CI [25.4, 43.8]) than during the dry season (9.1%, 95% CI [4.9, 16.5]), with the highest prevalence estimated on lettuce. Coliform levels on lettuce and tomatoes were greater during the rainy season (6.3 and 5.3 log CFU/g, 95% CI [5.7, 6.8] and [4.7, 5.8], respectively) than during the dry season (5.2 and 3.9 log CFU/g, 95% CI [4.7, 5.7] and [3.4, 4.4], respectively). These results indicate seasonal patterns for microbial prevalence in lettuce and an overall high level of bacterial contamination on raw vegetables sold in Cambodian informal markets.

Genotypic and Phenotypic Characterization of Antimicrobial Resistance Profiles in Non-typhoidal Salmonella enterica Strains Isolated From Cambodian Informal Markets.

Non-typhoidal Salmonella enterica is a pathogen of global importance, particularly in low and middle-income countries (LMICs). The presence of antimicrobial resistant (AMR) strains in market environments poses a serious health threat to consumers. In this study we identified and characterized the genotypic and phenotypic AMR profiles of 81 environmental S. enterica strains isolated from samples from informal markets in Cambodia in 2018-2019. AMR genotypes were retrieved from the NCBI Pathogen Detection website (https://www.ncbi.nlm.nih.gov/pathogens/) and using ResFinder (https://cge.cbs.dtu.dk/services/) Salmonella pathogenicity islands (SPIs) were identified with SPIFinder (https://cge.cbs.dtu.dk/services/). Susceptibility testing was performed by broth microdilution according to the Clinical and Laboratory Standards Institute (CLSI) standard guidelines M100-S22 using the National Antimicrobial Resistance Monitoring System (NARMS) Sensititre Gram Negative plate. A total of 17 unique AMR genes were detected in 53% (43/81) of the isolates, including those encoding tetracycline, beta-lactam, sulfonamide, quinolone, aminoglycoside, phenicol, and trimethoprim resistance. A total of 10 SPIs (SPI-1, 3-5, 8, 9, 12-14, and centisome 63 [C63PI]) were detected in 59 isolates. C63PI, an iron transport system in SPI-1, was observed in 56% of the isolates (n = 46). SPI-1, SPI-4, and SPI-9 were present in 13, 2, and 5% of the isolates, respectively. The most common phenotypic resistances were observed to tetracycline (47%; n = 38), ampicillin (37%; n = 30), streptomycin (20%; n = 16), chloramphenicol (17%; n = 14), and trimethoprim-sulfamethoxazole (16%; n = 13). This study contributes to understanding the AMR genes present in S. enterica isolates from informal markets in Cambodia, as well as support domestic epidemiological investigations of multidrug resistance (MDR) profiles.

Draft Genome Sequences of 81 Salmonella enterica Strains from Informal Markets in Cambodia.

Salmonella enterica is an important global pathogen due to its contribution to human morbidity and death. The presence of S. enterica in Southeast Asian informal markets is amplified by cross-contamination between market surfaces and food products. Here, we describe the draft genome sequences of 81 Salmonella enterica isolates from informal markets in Cambodia.