Faecal carriage of Verocytotoxin-producing Escherichia coli O157 and carcass contamination in cattle at slaughter in northern Italy.

A study on the prevalence of the faecal carriage of Verocytotoxin (VT)-producing Escherichia coli (VTEC) O157 and on the rate of carcass contamination was carried out on feedlot cattle and dairy cows at slaughter in northern Italy. Between April 1998 and January 1999, 12 sampling visits were performed on different days in seven different slaughterhouses. At each visit, 5-12 animals consecutively slaughtered were selected. From each animal, faeces were collected from the rectum immediately after slaughter and surface swabs were taken from the leg region and the diaphragmatic insertion of the carcass. All samples were examined for the presence of VTEC O157 using an immunomagnetic separation technique. A total of 100 animals coming from 60 different farms were examined. In total, VTEC O157 was isolated from the intestinal content of 17, and from the carcasses of 12 of the 100 animals examined. In particular, VTEC O157 was recovered from six (35.3%) out of the 17 carcasses from which the organism had previously been isolated from rectal content and from six (7.3%) of the 82 carcasses of the stool-negative cattle. In seven carcasses, VTEC O157 was isolated from the leg area, in two carcasses from the diaphragmatic area, and in three carcasses from both areas. Major differences in the prevalence of VTEC O157 were observed in the different groups of cattle sampled. In 7 of the 12 sampling visits, all the specimens examined were negative, while 16 of the 17 positive stool samples and 11 of the 12 positive carcass swabs were collected during three of the visits, performed in June in three different abattoirs. In these three visits, the ratios between the percentage of animals carrying VTEC O157 in the stools and the percentage of contaminated carcasses were 0.33, 0.57, and 1.66, respectively; thus, confirming that slaughter practices can largely influence the rate of carcass contamination. Phage typing and PFGE analysis of VTEC O157 isolated from samples collected at the same visit suggested that both auto- and cross-contamination occurred.     

Detection of Escherichia coli O157 in bovine meat products in northern Italy

Tests for Escherichia coli and E. coli O157 were carried out on meat samples collected from randomly chosen stores throughout the city of Bologna and suburban areas. The samples consisted of 25 g of loose minced beef, sometimes already shaped into meatballs or hamburgers, some of which were mixed with vegetables. The meat was purchased from retail outlets, open market stalls, and supermarket chains during 25 sampling visits from October 2000 to December 2001. For E. coli detection, Tryptone soya broth (TSB) supplemented with novobiocin and C-EC agar were used. Immunomagnetic separation with SMAC-BCIG-CT agar and chromogenic E. coli O 157 agar, API 20E system and agglutination latex test were used to detect E. coli O157; Vero cell assay and polymerase chain reaction (PCR) were used to assess toxin production and the presence of virulence genes.

E. coli were detected in 45 (30.2%) of the 149 samples examined, mainly in the hamburger samples mixed with vegetables and in the loose minced beef. E. coli O157 was found in one sample of hamburger and two samples of hamburger mixed with vegetables (2%) collected from three different butcher's stores between July and October. All the strains of E. coli O157 and most cases of E. coli were found in meat from small retailers.

The three strains of E. coli O157 were positive for verocytotoxin production. PCR analysis revealed genes coding for vt2 and one strain possessed the gene for eae A. Chromogenic E. coli O157 agar was found to be more selective and differential, allowing easier identification of suspected colonies with mixed flora and producing less false-positive colonies.     

Escherichia coli O26 in minced beef: prevalence, characterization and antimicrobial resistance pattern

Verocytotoxin-producing Escherichia coli (VTEC) non-O157 serogroups are among the most important emerging food-borne pathogen groups. In particular, the O26 serogroup is able to cause a large spectrum of illnesses in humans which have a significant public health impact as they may range from haemorrhagic colitis (HC) to haemolytic uremic syndrome (HUS) and thrombotic thrombocytopenic purpura (TTP). It is known that VTEC organisms are associated with animal reservoirs, i.e. ruminants, and foods of animal origin, especially undercooked meat and raw milk, are often involved in outbreaks. In this study, 250 minced beef samples collected at retail outlets in southern Italy were tested for the presence of E. coli O26 and the isolates were characterized and studied for their antimicrobial resistance properties. Three minced beef samples (1.2%) tested positive for E. coli O26; one isolate per positive sample was characterized. One isolate harboured the genes encoding for virulence factors intimin (eaeA) and enterohaemolysin (hlyA), while none presented verocytotoxin-encoding genes (stx1 and stx2) and all were negative at the verotoxicity assay. All the isolates showed resistance properties to at least four antimicrobial agents tested and two were multi-drug resistant (MDR). Although no verocytotoxin-encoding genes were found in the isolates, the presence of potentially pathogenic E. coli O26 strains in minced beef points to the need for proper hygiene during meat production to reduce the risk of food-borne illnesses and transmission of MDR organisms via foods to humans. This paper is the first report on the presence and characterization of E. coli O26 in minced beef marketed in Italy.     

Applicability of the draft standard method for the detection of Escherichia coli O157 in dairy products

According to the draft standard method, EN ISO 16654 for the detection of Escherichia coli O157 in foods, samples of milk products inoculated with E. coli O157 were cultured in modified Tryptone Soya Broth supplemented with novobiocin. After immuno-magnetic separation (IMS) of the micro-organisms with magnetic beads coated with an antibody against E. coli O157 (Dynabeads anti E. coli O157, Dynal), the enrichment broth was subcultured onto both Cefixim Tellurite Sorbitol MacConkey agar and CHROMagar O157. IMS alone appeared not to be very specific to E. coli O157; however, IMS combined with CT-SMAC plating enabled a greater exclusivity. The method displayed a low limit of detection (1-2 cfu/25 g) of E. coli O157 strains in milk products after only 6 h of incubation of the enrichment broth. However, the detection was affected by storage of the inoculum at 4 degrees C, and another 12 to 18 h of incubation was necessary to recover potentially stressed cells.     

Evaluation of increased incubation temperature and cefixime-tellurite treatment for the isolation of Escherichia coli O157:H7 from minced beef

To improve enrichment and isolation of Escherichia coli O157:H7, this study evaluated increased incubation temperature and cefixime-tellurite (CT) on five strains of each of the following bacteria, E. coli, Hafnia alvei, Enterobacter spp., Citrobacter freundii and E. coli O157:H7, and two strains of E. coli O157:nH7. These were grown in pure culture in LST broth with varying cefixime-tellurite concentrations. A range of incubation temperatures from 37 to 46 degrees C was investigated for the inhibition of cohabitant microorganisms. Minced beef, spiked with E. coli O157:H7 and cohabitant microorganisms was investigated. Increased incubation temperature (42 degrees C) and treatment with half of the prescribed amount of cefixime-tellurite by BAM for SMAC agar in enrichment step were the most effective in selectively growing E. coli O157:H7. The results show that E. coli O157:H7 is more resistant to these two conditions than the other cohabitant bacteria.     

Escherichia coli O157 prevalence and enumeration of aerobic bacteria, Enterobacteriaceae, and Escherichia coli O157 at various steps in commercial beef processing plants

The effectiveness of current antimicrobial interventions used in reducing the prevalence or load of Escherichia coli O157 and indicator organisms on cattle hides and carcasses at two commercial beef processing plants was evaluated. Sponge sampling of beef cattle was performed at five locations from the initial entry of the animals to the slaughter floor to the exit of carcasses from the "hotbox" cooler. For each sample, E. coli O157 prevalence was determined and total aerobic bacteria, Enterobacteriaceae, and E. coli O157 were enumerated. E. coli O157 was found on 76% of animal hides coming into the plants, but no carcasses leaving the cooler were identified as contaminated with E. coli O157. A positive relationship was seen between the incidence of E. coli O157 in hide samples and that in preevisceration samples. Aerobic plate counts and Enterobacteriaceae counts averaged 7.8 and 6.2 log CFU/100 cm2, respectively, on hides, and 1.4 and 0.4 log CFU/100 cm2, respectively, on chilled carcasses. Aerobic plate counts and Enterobacteriaceae counts on preevisceration carcasses were significantly related to the respective levels on the corresponding hides; the carcasses of animals whose hides carried higher numbers of bacteria were more likely to carry higher numbers of bacteria. Implementation of the sampling protocol described here would allow processors to evaluate the efficacy of on-line antimicrobial interventions and allow industrywide benchmarking of hygienic practices.     

Presence of Escherichia coli O157:H7 in ground beef and ground baby beef meat

A total of 114 beef and baby beef samples were examined. The samples included ground baby beef, mixed ground baby beef and pork, and chopped and shaped meat. The samples were analyzed from 30 different grocery stores in Zagreb, Croatia. The object of this study was to evaluate the prevalence of Escherichia coli O157:H7 in the samples that can enhance the potential risk of outbreaks of hemorrhagic colitis and hemolytic uremic syndrome. The results in all tested samples of E. coli O157:H7 were negative. A single sample was positive in a latex agglutination test using antiserum to O157:H7. It was identified as Proteus vulgaris at the Pasteur Institute, Paris, France. This result correlates positively with cross-contamination with Yersinia enterocolitica 09, Brucella abortus, Salmonella type N, and Pseudomonas maltophila.     

Fate of field-isolated Escherichia coli O157 in ground beef at different storage temperatures

The survival of six Escherichia coli O157 strains, including five strains recently isolated from beef carcasses and strain ATCC 43895, was evaluated at 0, 1, 7, and 14 days in ground beef held at -20, 1, 4, and 7 degrees C. Only small losses in cell numbers occurred at -20 and 1 degree C; in general, cell numbers decreased during the first day of storage and then remained unchanged through day 14. At -20 degrees C, statistically significant reductions in cell numbers were observed only for strains 55AC1 and 299AB3 due to greater losses in the first day. At 1 degree C, strain 131AC1 did not decrease in cell numbers during the first day of storage, but both this strain and strain 55AC1 experienced statistically significant reductions in viable cell numbers by day 14, primarily due to losses after day 7. At 4 degrees C, after an initial loss of cell numbers for four strains, minor increases were observed for all six strains by day 14. The differences were statistically significant for strains 114AC1, 299AB3, and ATCC 43895, but were small enough to question whether they refect actual growth. When the inoculated ground beef was stored at 7 degrees C for 14 days, growth of all six strains was statistically significant, with populations increasing between 0.9 and 1.5 log10 CFU/g. This study demonstrates that there are small differences in the abilities of various E. coli O157 strains to survive and sometimes grow in fresh ground beef at cold storage temperatures, but overall these differences do not appear to be meaningful. The differences cannot be attributed to recency of isolation, since strain ATCC 43895 behaved similarly to recently isolated strains. Storage temperatures of 4 degrees C or below limited growth of E. coli O157 isolates, but did not have a noteworthy effect on survival.     

Seasonal prevalence of Escherichia coli O157:H7 in beef cattle feces

Cattle are an asymptomatic reservoir of Escherichia coli O157:H7, but the bacterial colonization and shedding patterns are poorly understood. The prevalence and shedding of this human pathogen have been reported to be seasonal with rates typically increasing during warm months. The objectives of this study were (i) to assess the prevalence of E. coli O157:H7 in feces of feedlot cattle in Kansas during summer, fall, and winter months, and (ii) to characterize E. coli O157:H7 by screening for virulence factors. Of 891 fecal samples collected, 82 (9.2%) were positive for E. coli O157:H7. No significant differences in prevalence were detected among summer, fall, and winter months. The highest monthly prevalence (18.1%) was detected in February. All tested isolates were positive for stx2 (Shiga toxin 2) and eaeA (intimin) genes; 14 isolates (12.8%) also carried stx1. Our results indicate the prevalence of E. coli O157:H7 in beef cattle feces is not necessarily season dependent.     

Process control and sampling for Escherichia coli O157:H7 in beef trimmings

Raw beef producers currently face the problem of Escherichia coli O157:H7 surface contamination of beef carcasses that can lead to product adulteration. Although carcass interventions are in place, elimination of E. coli O157:H7 from every potential hiding place on the surfaces of a beef carcass is not technologically feasible. Therefore, E. coli O157:H7 on beef carcasses might further contaminate the surfaces of beef trimmings. With the use of case scenarios from nine commercial processing facilities, we present a process control and statistical sampling approach for monitoring beef trimmings to divert contaminated lots of the trimmings from the raw ground beef supply chain.     

Escherichia coli O157:H7 shedding in vaccinated beef calves born to cows vaccinated prepartum with Escherichia coli O157:H7 SRP vaccine

Extensive research, intervention equipment, money, and media coverage have been directed at controlling Escherichia coli O157:H7 in beef cattle. However, much of the focus has been on controlling this pathogen postcolonization. This study was conducted to examine the performance, health, and shedding characteristics of beef calves that were vaccinated with an E. coli O157:H7 SRP bacterial extract. These calves had been born to cows vaccinated prepartum with the same vaccine. Cows and calves were assigned randomly to one of four treatments: (i) neither cows nor calves vaccinated with E. coli O157:H7 SRP (CON), (ii) cows vaccinated with E. coli O157:H7 SRP prepartum but calves not vaccinated (COWVAC), (iii) calves vaccinated with E. coli O157:H7 SRP but born to cows not vaccinated (CALFVAC), (iv) cows vaccinated with E. coli O157:H7 SRP prepartum and calves also vaccinated (BOTH). Calves born to vaccinated cows had significantly higher titers of anti-E. coli O157:H7 SRP antibodies (SRPAb) in circulation at branding time (P < 0.001). Upon entry to the feedlot, overall fecal E. coli O157:H7 prevalence was 23 % among calves, with 25 % in the CON treatment group, 19 % in the CALFVAC group, 32 % in the COWVAC group, and 15 % in the BOTH group (P > 0.05). Fecal shedding of E. coli O157 on arrival to the feedlot was not correlated with fecal shedding at slaughter (Spearman's rho = -0.02; P = 0.91). No significant effects of cow or calf E. coli O157:H7 SRP vaccination treatment were found on feedlot calf health or performance (P > 0.05), prevalence of lung lesions or liver abscess (P > 0.05), or morbidity, retreatment, or mortality numbers (P > 0.05). The findings of this study indicate that the timing of vaccination of calves against E. coli O157:H7 may be an important consideration for maximizing the field efficacy of this vaccine.     

The antimicrobial effect of thyme essential oil, nisin and their combination against Escherichia coli O157:H7 in minced beef during refrigerated storage

The antimicrobial effect of thyme essential oil (EO) at supplementation levels of 0.3%, 0.6% or 0.9%, nisin at 500 or 1000 IU/g, and their combination, on Escherichia coli O157:H7 was examined in both tryptic soy broth (TSB) and minced beef meat. EO at 0.3% possessed a weak antibacterial activity against the pathogen in TSB, whereas at 0.9% showed unacceptable organoleptic properties in minced meat. Thus, only the level of 0.6% of EO was further examined against the pathogens in minced meat. Treatment of minced beef meat with EO at 0.6% showed an inhibitory activity against E. coli O157:H7 during storage at 10 °C, but not at 4 °C. Treatment of minced beef meat or TSB with nisin at 500 or 1000 IU/g did not show any antibacterial activity against E. coli O157:H7. The combination of EO at 0.6% and nisin at 500 or 1000 IU/g showed an additive effect against the pathogen, which was higher during storage at 10 °C than at 4 °C.     

An evaluation of rapid methods for detecting Escherichia coli O157 on beef carcasses

Numbers of Escherichia coli O157 in food may be low and sensitive techniques are therefore needed for its detection. The objectives of this study were to use carcass meat samples artificially inoculated with various strains of E. coli O157 to compare the sensitivity of enrichment in three different media and to compare immunomagnetic separation followed by culture of magnetic beads to cefixime tellurite sorbitol MacConkey agar with three immunoassays for the detection of E. coli O157 in the enrichment cultures. Duplicate 250, 25 and 2-3 CFU of each of 16 strains of E. coli O157 added to 25-g samples of beef carcass meat were used to compare the sensitivity of (1) enrichment in supplemented tryptone soya broth (sTSB), Reveal 8-h and Reveal 20-h media, and (2) immunomagnetic separation and culture to cefixime tellurite sorbitol MacConkey agar (IMS/CT-SMAC) with Reveal, VIP and STAT immunoassays for detecting the organism. An initial inoculum of 250 CFU/25 g meat was detected in all 32 samples by IMS/CT-SMAC performed on all enrichment media and by Reveal performed on Reveal 8-h and Reveal 20-h media, but in only 30, 19 and 9 samples by Reveal, VIP and STAT, respectively, performed on sTSB medium. An initial inoculum of 25 CFU/25 g meat was detected in 28, 32 and 30 of 32 samples by IMS/CT-SMAC performed on sTSB, Reveal 8-h and Reveal 20-h media, respectively, and in 32 and 30 samples by Reveal performed on Reveal 8-h and Reveal 20-h media, but in only 22, 11 and 2 samples by Reveal, VIP and STAT, respectively, performed on sTSB medium. An initial inoculum of 2-3 CFU/25 g meat was detected in 25, 31 and 28 of 32 samples by IMS/CT-SMAC performed on sTSB, Reveal 8-h and Reveal 20-h media, respectively, and in 25 and 23 samples by Reveal performed on Reveal 8-h and Reveal 20-h media, but in only 14, 1 and 0 samples by Reveal, VIP and STAT, respectively, performed on sTSB medium.     

Growth rates of Salmonella and Escherichia coli O157:H7 in irradiated beef.

Ground beef was irradiated to 0, 2, or 4 kGy and then inoculated with a mixed culture of four serotypes of salmonellae or five strains of Escherichia coli O157:H7. The ground beef was stored at either 15 or 25 degrees C, and the growth of the inoculated bacteria was monitored over time. Growth parameters were determined for both the salmonellae and the E. coli O157:H7 using the Gompertz equation. There was no significant difference in lag phase duration or generation time, irrespective of the dose to which the ground beef had previously been exposed. Furthermore, the lag phase durations and generation times determined in this study did not differ significantly from previously published values. This suggests that, although irradiation eliminates a significant portion of the spoilage microflora in ground beef, the absence of this microflora provides no competitive advantage to the growth of salmonellae or E. coli O157:H7 in ground beef.     

Modeling preharvest and harvest interventions for Escherichia coli O157 contamination of beef cattle carcasses

Field studies evaluating the effects of multiple concurrent preharvest interventions for Escherichia coli O157 are logistically and economically challenging; however, modeling techniques may provide useful information on these effects while also identifying crucial information gaps that can guide future research. We constructed a risk assessment model with data obtained from a systematic search of scientific literature. Parameter distributions were incorporated into a stochastic Monte Carlo modeling framework to examine the impacts of different combinations of preharvest and harvest interventions for E. coli O157 on the risk of beef carcass contamination. We estimated the risk of E. coli O157 carcass contamination conditional on preharvest fecal prevalence estimates, inclusion of feed additive(s) in the diet, vaccination for E. coli O157, transport and lairage effects, hide intervention(s), and carcass intervention(s). Prevalence parameters for E. coli O157 were assumed to encompass potential effects of concentration; therefore, concentration effects were not specifically evaluated in this study. Sensitivity analyses revealed that fecal prevalence, fecal-to-hide transfer, hide-to-carcass transfer, and carcass intervention efficacy significantly affected the risk of carcass contamination (correlation coefficients of 0.37, 0.56, 0.58, and -0.29, respectively). The results indicated that combinations of preharvest interventions may be particularly important for supplementing harvest interventions during periods of higher variability in fecal shedding prevalence (i.e., summer). Further assessments of the relationships among fecal prevalence and concentration, hide contamination, and subsequent carcass contamination are needed to further define risks and intervention impacts for E. coli O157 contamination of beef.     

Escherichia coli O157 in cattle and sheep at slaughter, on beef and lamb carcasses and in raw beef and lamb products in South Yorkshire, UK

A 1 year study of Escherichia coli O157 in cattle and sheep at slaughter, on beef and lamb carcasses and in raw beef and lamb products from retail butchers' shops was performed in the Sheffield area. Each month, samples of rectal faeces were collected immediately after slaughter from 400 cattle and 600 sheep, and 400-430 samples of raw meat products were purchased from butchers' shops. Meat samples were also obtained from 1500 beef and 1500 lamb carcasses. All samples were examined for E. coli O157 by enrichment culture, immunomagnetic separation and culture of magnetic particles onto cefixime tellurite sorbitol MacConkey agar. Raw meat products were also examined for numbers of generic E. coli by a standard membrane culture method. E. coli O157 was isolated from 620 (12.9%) of 4800 cattle, 100 (7.4%) of 7200 sheep, 21 (1.4%) of 1500 beef carcasses, 10 (0.7%) of 1500 lamb carcasses and from 22 (0.44%) of 4983 raw meat products. E. coli O157 was isolated more frequently from lamb products (0.8%) than from beef products (0.4%). Numbers of generic E. coli in meat products reached seasonal peaks in July and August with counts of > 10(4)/g occurring more frequently in lamb products (50.8 and 42.4%, respectively) than in beef products (19.3 and 23.8%, respectively). The majority of E. coli O157 strains, from animals, carcasses and meat samples, were isolated during the summer. Most were verocytotoxigenic as determined by Vero cell assay and DNA hybridisation, eaeA gene positive and contained a 92 kb plasmid. The isolates were compared with 66 isolates from human cases over the same period. A combination of phage type, toxin genotype and plasmid analysis allowed subdivision of all the E. coli O157 isolates into 96 subtypes. Of these subtypes, 53 (55%) were isolated only from bovine faecal samples. However, 61 (92%) of the 66 isolates from humans belonged to 13 subtypes which were also found in the animal population.     

Occurrence of Shiga toxin-producing Escherichia coli O157 in selected dairy and meat products marketed in the city of Rabat, Morocco

Samples of meat and dairy products taken from the city of Rabat, Morocco, were examined for the presence of Escherichia coli O157 by the selective enrichment procedure followed by plating on cefixime-tellurite-sorbitol MacConkey agar and a latex agglutination test. The ability of isolates to produce Shiga toxins (ST1 or ST2) was also tested by an agglutination test using sensitized latex. Dairy samples (n = 44) included different products commonly consumed in the country. Meat samples (n = 36) were taken from traditional butchers because these products are generally marketed in this way. Random samples were taken from each product during the period of January through May. Of the 80 samples tested, 8 (10%) harbored E. coli O157. Four dairy and four meat samples were contaminated (9.1 and 11.1%, respectively). Of 10 E. coli O157 isolates from contaminated samples demonstrating true antigen-antibody agglutination, 5 (50%) produced either ST2 alone or ST2 plus ST1. Four of the five strains (80%) were meat isolates and produced ST2 with or without ST1, and the fifth was a dairy isolate producing ST2.     

Thermal process validation for Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes in ground turkey and beef products

At 55 to 70 degrees C, thermal inactivation D-values for Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes were 19.05 to 0.038, 43.10 to 0.096, and 33.11 to 0.12 min, respectively, in ground turkey and 21.55 to 0.055, 37.04 to 0.066, and 36.90 to 0.063 min, respectively, in ground beef. The z-values were 5.73, 5.54, and 6.13 degrees C, respectively, in ground turkey and 5.43, 5.74, and 6.01 degrees C, respectively, in ground beef. In both ground turkey and beef, significant (P < 0.05) differences were found in the D-values between E. coli O157:H7 and Salmonella or between E. coli O157:H7 and L. monocytogenes. At 65 to 70 degrees C, D-values for E. coli O157:H7, Salmonella, and L. monocytogenes were also significantly (P < 0.05) different between turkey and beef. The obtained D- and z-values were used in predicting process lethality of the pathogens in ground turkey and beef patties cooked in an air impingement oven and confirmed by inoculation studies for a 7-log (CFU/g) reduction of E. coli O157:H7, Salmonella, and L. monocytogenes.     

Prevalence and antibiotic resistance profiles of Escherichia coli O157:H7 in beef products from retail outlets in Gaborone, Botswana

Four hundred meat samples (134 meat cubes, 133 minced meat, 133 fresh sausages) were collected from 15 supermarkets and butcheries in Gaborone, Botswana, between the summer months of October 2002 and March 2003. Samples were assayed for Escherichia coli O157 by selective enrichment in modified E. coli broth containing novobiocin, followed by immunomagnetic separation and plating onto sorbitol MacConkey agar supplemented with potassium tellurite. The isolates were biochemically and serologically confirmed by API 20E and O157 antisera, respectively. The prevalence rates for E. coli O157 were 5.22% in meat cube samples, 3.76% in minced meat samples, and 2.26% in fresh sausages. The isolates showed single, double, and triple antibiotic resistance. Fifty-three percent of them were resistant to cephalothin. Resistance was also recorded for sulphatriad (33%), colistin sulphate (26%), streptomycin (0.7%), and tetracycline (26%). It is recommended that the cause for antibiotic resistance be investigated using a larger number of samples from cattle, especially from ranching areas of the country.