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. 2009 Oct;6(8):965–972. doi: 10.1089/fpd.2008.0239

Salmonella enterica Subclinical Infection: Bacteriological, Serological, Pulsed-Field Gel Electrophoresis, and Antimicrobial Resistance Profiles—Longitudinal Study in a Three-Site Farrow-to-Finish Farm

German B Vigo 1, Javier A Cappuccio 2, Pablo E Piñeyro 2,,3,✉, Angela Salve 4, Mariana A Machuca 2, Maria A Quiroga 2, Fabiana Moredo 1, Gabriel Giacoboni 1, Jose L Cancer 5, Ines G Caffer 4, Norma Binsztein 4, Mariana Pichel 4, Carlos J Perfumo 2
PMCID: PMC3145166  PMID: 19642916

Abstract

The aim of this surveillance was to study both Salmonella spp. shedding patterns and the time course of serological response in farrow-to-finish reared pigs from a subclinically infected farm. Antimicrobial resistance profile, molecular subtyping, and the relationship among the isolates were determined by pulsed-field gel electrophoresis (PFGE). A farrow-to-finish farm of 6000 sows, with a history of Salmonella Typhimurium septicemia, was selected. A longitudinal bacteriological and serological study was conducted in 25 sows before farrowing (M/S1) and in 50 offspring at 21 (M/S2), 35 (M/S3), 65 (M/S4), 86 (M/S5), 128 (M/S6), and 165 (M/S7) days of age. Serum antibodies were tested using Herdcheck® Swine Salmonella antibody test kit (Idexx Laboratories, ME). Bacteria were isolated from pooled fecal samples. Suspected isolates were confirmed by conventional biochemical assays, and those identified as Salmonella spp. were serotyped. A variation between seropositive percentages and positive fecal samples was observed. Serologically positive pigs decreased from S1 to S4, and subsequently increased from S4 to S7. The percentages of fecal positive culture increased from M1 to M3, and then declined in M4, increased in M5, and were negative in M6 and M7.

In the study three serovars, Salmonella 3,10:e,h:-, Salmonella Muenster, and Salmonella Bovismorbificans, were identified with low pathogenicity for swine. Three multidrug resistance strains (one belonged to Salmonella 3,10:e,h:- and two belonged to Salmonella Muenster) were found. PFGE results showed three different but closely related patterns among the 13 isolates of Salmonella Bovismorbificans, and two patterns for the three Salmonella Muenster and Salmonella 3,10:e,h:- isolates.

This longitudinal study established critical points of Salmonella spp. infection in the farm and the production stages, where appropriate control measures must be taken. PFGE showed clonal relationships in each serovar. Antibiotic resistance profiles should be periodically included due to public health concerns.

Introduction

Salmonella spp. infection is of major concern to the swine industry for several important reasons: (1) contaminated pork distributed in retail stores can present very significant risk to public health, and (2) septicemic salmonellosis, due to host-adapted Salmonella cholerasuis, or severe enteritis, due to nonhost-adapted Salmonella Typhimurium (Lanza, 1998; Weiss et al., 2002; Griffith et al., 2006), causes significant mortality in piglet and grower pigs, resulting in important economic loss.

Salmonella spp. are one of the most important foodborne pathogens transmitted to humans worldwide (Asai et al., 2002; Beloeil et al., 2003; Oliveira et al., 2005). Pork is the second most important source of Salmonella spp. in some countries, and many human outbreaks reported in the last decade have been traced to consumption of pork meat (Griffith et al., 2006).

It is assumed that the greater the number of Salmonella spp. that are carried into the slaughter process, via the pigs' intestines, the greater the risk of equipment and final product contamination (Hurd et al., 2002; Sauli et al., 2005; Griffith et al., 2006). Therefore, reductions in preslaughter infection rates should result in increased pork safety (Hurd et al., 2002; Oliveira et al., 2005; Sauli et al., 2005).

Infection of swine by one or more serovars is common (Griffith et al., 2006), but nowadays Salmonella spp. do not usually cause clinical disease in swine (Kranker et al., 2003). Enterocolitis due to Salmonella Typhimurium occurs more frequently than expected in high health herds (Griffith et al., 2006); however, this serovar can also live on swine intestines without clinical signs (Griffith et al., 2006). Salmonella spp. are frequently isolated as a sequel to other enteric or debilitating diseases such as porcine circovirus 2 (Griffith et al., 2006; Silva et al., 2006) and other predisposing factors (Schawrtz, 1997; Vigo et al., 2004; Ha et al., 2005).

Generally, to estimate Salmonella spp. prevalence in pig herds, bacteriological examinations are performed on fecal samples or tissues from clinically affected animals (Asai et al., 2002). Culturing fecal samples for Salmonella spp. is a useful tool to determine current infections in a pig herd. However, Salmonella spp. shedding by pigs is intermittent with a low number of bacteria in the feces of subclinically infected pigs (Oliveira et al., 2005). Therefore, conventional culture methods are labor intensive, time consuming, and expensive, and may not be practical or economically feasible for routine application due to low sensitivity (Ekeroth et al., 2003; Lo Fo Wong et al., 2003). Modern serological techniques, such as enzyme-linked immunosorbent assay tests, have proven to be convenient and cost-effective methods for screening for antibodies against Salmonella spp. (Lo Fo Wong et al., 2003). Measuring antibody responses is an indirect but more sensitive method to determine the prevalence of pigs harboring the bacteria in a herd (Ekeroth et al., 2003). Importantly, the presence of serum antibodies reflects previous exposure rather than current infection (Funk et al., 2005). Neither fecal culture nor detection of serum antibodies represents a perfect diagnostic test for Salmonella spp. Apart from this, the limitations of both tests should be considered to interpret the results (Nollet et al., 2005b).

To determine the dynamics of Salmonella spp. infection in swine herds over time (e.g., age, duration of infection, and disease transmission patterns), longitudinal studies following the bacteriological and serological status of pigs should be performed (Kranker et al., 2003; Oliveira et al., 2005). Further, the distribution of Salmonella spp. and its clonal relationships between strain types circulating among different farm facilities provide valuable information to understand the epidemiology of the infection (Zhao et al., 2007).

Approaches to prevent and control salmonellosis in the food animal industry include, among others, improved biosecurity, vaccination, and use of competitive exclusion products, although each of these practices has had limited success. The control and prevention of Salmonella spp. needs to be reinforced by the use of antimicrobial chemotherapy (Zhao et al., 2007). Data show increased antimicrobial resistance among several Salmonella serovars, and multiple drug resistance (MDR) is an emerging problem (Douris et al., 2008). The antimicrobial resistance makes it more difficult for clinicians to empirically select an appropriate antibiotic (Zhao et al., 2007).

Conventional and molecular epidemiology and antimicrobial susceptibility profiles are important epidemiological tools to determine potential sources of infections and have been widely used to understand the epidemiology of many infectious diseases (Liebana, 2002). The current gold standard method of choice for molecular typing of Salmonella spp. as a source identification is pulsed-field gel electrophoresis (PFGE) (Ridley et al., 1998; Liebana, 2002; Best et al., 2007; Vigo et al., 2007).

Currently, there is limited published data relative to the prevalence, antimicrobial susceptibility, and molecular subtypes of Salmonella spp. present in swine herds in Argentina (Vigo et al., 2004, 2007). The aim of this survey was to study both the Salmonella spp. shedding patterns and the time course of the serological response in farrow-to-finish reared pigs from a subclinically infected farm, and to determine the antimicrobial resistance profile, molecular subtype by PFGE, and the relationship among the isolates.

Materials and Methods

Farm selection and sampling size

A farrow-to-finish farm (6000 sows) was selected. The farm consisted of three operation units and used a batch farrowing system (250 sows/week) with an all-in/all-out hygiene policy at farrowing, postweaning, and fattening sections. The piglets were weaned at 3 weeks and moved to site 2. Site 2, where growers are raised, has nine barns and all of them are loaded with piglets weaned for a full week (3000 piglets). After 8 weeks in site 2, growers are moved to site 3, which has 14 barns and allows raising 1500 finishers in each one of them, until they reach the market weight.

Salmonella spp. positive status of the farm was confirmed by a preliminary study carried out in October 2005, when an increased mortality in fattening units was observed, and Salmonella Typhimurium was isolated from the spleen of pigs that suffered from porcine dermatitis and nephropathy syndrome (Vigo et al., 2004).

Sampling scheme

Site 1 (farrowing)

At the beginning of this study, 25 sows, within a weekly batch, were randomly selected (Fig. 1). They were bled the day before farrow (S1). The day after farrow, 10–25 g of feces was collected from the rectum of sows with sterile gloves or from the floor using sterile plastic bags (M1). From 10 of the previously selected sows, a total of 50 piglets were randomly selected and tagged. Piglets were bled at weaning age (21 ± 3 days old) (S2). At the same time, fecal samples, 10 pools of 10–25 g, each pool coming from five animals, were collected from the floor of the farrowing crates (M2).

FIG. 1.

FIG. 1.

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Longitudinal sampling scheme for serological and bacteriological studies. M, fecal material for bacteriological studies; S, blood sample for serological studies. The samples of M1 and S1 corresponded with 25 randomly selected sows sampled 1 day prefarrowing (M1) and 1 day postfarrowing (S1), respectively. The samples from M2 to M6 and S2 to S6 corresponded with those obtained for bacteriological and serological studies, respectively, in each stage of production in the cohort of studied piglets.

Site 2 (nursery)

The 50 pigs previously tagged were sampled 2 weeks after their transfer to site 2 (35 ± 3 days old). Individual blood samples (S3) and pooled fecal samples for bacteriological studies were collected (M3). They were taken from the floor in five different points of each pen where the piglets were held. The same sample scheme for serological (S4) and bacteriological (M4) studies was carried out 1 week before the pigs left site 2 (65 ± 3 days old).

Site 3 (fattening)

In this stage, three samples for serological and bacteriological studies were taken as described above. The first sample was collected 2 weeks after the pigs reached site 3 (86 ± 3 days old) (S5) (M5), the second sample was collected 8 weeks into the finisher stage (midway through the fattening period, 128 ± 3 days old) (S6) (M6), and the last sample was taken 1 week before the pigs left the farm for slaughter (165 ± 3 days old) (S7) (M7).

Serology

Blood was taken from the cava cranialis vein. The samples were centrifuged the same day, and sera were collected and stored at −20°C until study. Serum antibodies were tested using Herdcheck Swine Salmonella antibody test kit (Idexx Laboratories) following the manufacturer's instructions. Optical densities (ODs) were determined by a spectrophotometer with a 650 nm filter. The test detects the most common serogroups (B, C1, and D) isolated in Europe, Asia, and America, and has a specificity of 99.4% (HerdChek Swine Salmonella Information Sheet; Idexx Laboratories).

The ODs were calculated as

graphic file with name M1.gif

Samples with OD ≥0.4 were considered to be positive. Software xChek™ 3.3 (Idexx Laboratories) was used to obtain the results.

Bacteriology

From each pooled sample, 10–25 g of feces was diluted 1/10 in buffered peptone water. Each sample was incubated for 24 h at 37°C (preenrichment). Thereafter, 1 mL of broth was inoculated in 10 mL of tetrathionate broth for 24 h at 37°C (enrichment). A loopful of enrichment broth was transferred to enteric Hektoen agar with novobiocin (10 μg/mL) and incubated for 48 h at 37°C. Two presumptively positive colonies were streaked in tripticase soy agar and incubated for 24 h at 37°C, and were subsequently confirmed by biochemical conventional assays (Koneman et al., 1999). Isolates identified as Salmonella spp. were serotyped by agglutination according to M. Poppof's (Clinical and Laboratory Standards Institute, 2005) scheme, using polyvalent and monovalent O and H antisera produced at the Servicio de Antígenos y Antisueros, Instituto Nacional de Producción de Reactivos y Biológicos—ANLIS “Carlos G. Malbrán.”

Antimicrobial resistance assay

Susceptibility of Salmonella spp. strains to ampicillin, cephalothin, cefotaxime, norfloxacin, enrofloxacin, nalidixic acid, gentamicin, streptomycin, amikacin, chloramphenicol, fosfomycin, polimixin, tetracycline, nitrofurantoin, and trimethoprim-sulphamethoxazole was established by disk diffusion according to the Clinical Laboratory Standards Institute (Hunter et al., 2005).

PFGE

This was carried out following PulseNet standardized protocol from the Centers for Disease Control and Prevention (Ribot et al., 2006). Briefly, the plugs of agarose containing DNA were digested with 30 U of XbaI (Promega, Madison, WI). Fragments were separated in a 1% agarose gel (Seakem Gold; Lonza, Rockland, ME) in 0. 5 × Tris borate EDTA buffer at 14°C in a contour CHEF-DR III System (Bio-Rad, Hercules, CA). Run time was 18 h, with a constant voltage of 200 V, using linear ramp of 2.2–63.8 sec. Staining was carried out with 0.5 μg/mL of aqueous ethidium bromide solution (Bio-Rad). Salmonella Braenderup CDC-H-9812 was included as fragment size marker to analyze the patterns generated with XbaI-PFGE (Funk et al., 2001). The images of PFGE gels were obtained by Gel-Doc 2000 System (Bio-Rad), and were analyzed using BioNumerics version 3.5 (Applied Maths, Kortrij, Belgium). The relationship among the patterns was estimated as the proportion of shared bands applying the Dice coefficient with a 1.5% band position tolerance, and a dendrogram based on the unweighted pair group method with the arithmetic mean method was generated.

Results

At the beginning of the study, eight of the pigs tagged died of causes not related with Salmonella spp. infection.

Figure 2 shows the temporal variation of seropositive pigs and positive fecal samples found throughout the study. Briefly, percentage of positive sera clearly decreased from S1 to S4, and subsequently increased from S4 to S7. The percentage of fecal Salmonella-positive cultures increased from M1 to M3, and then abruptly declined in M4 and increased in M5; all the samples were negative in M6 and M7. During the study, only three Salmonella serovars were identified (Table 1).

FIG. 2.

FIG. 2.

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The bacteriological (M1) and serological (S1) status of the samples throughout the production. Percentage of Salmonella-positive samples detected (open bars, bacteriology, M2–M7) and percentages of seropositive animal (shaded bars, serology, S2–S7) are shown. Error bars indicate 95% confidence interval.

Table 1.

Salmonella Serovars and Resistance Profiles of the Isolates Recovered in Each Stage Sampled

Sample n Number of isolates Salmonella serovars Resistance profile
M1 23 1 Salmonella 3,10:e,h:- Nit, Tet, Nal, Cmp
M2 11 1 Salmonella Muenster Nit, Tet, Nal
M3 10 8 Salmonella Bovismorbificans –
  1 Salmonella Muenster Nit, Tet  
M4 10 1 Salmonella Bovismorbificans –
M5 10 4 Salmonella Bovismorbificans –
  1 Salmonella Muenster Tet  
M6 10 0 Negative  
M7 10 0 Negative  
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Nit, nitrofurantoin; Tet, tetracycline; Nal, nalidixic acid; Cmp, chloramphenicol.

While Salmonella Bovismorbificans isolates were sensitive to all the antimicrobial agents assayed, Salmonella Muenster and Salmonella 3,10:e,h:- isolates showed resistance to at least one antibiotic (Table 1).

To determine the genetic relatedness among the isolates belonging to each serovar, strains were analyzed by PFGE. The subtyping results for the 13 Salmonella Bovismorbificans isolates showed that three XbaI-PFGE patterns were identified. These patterns showed very slight differences in the position of only one band of high molecular weight and were distinct from other Salmonella Bovismorbificans isolates present in the Salmonella PFGE National Database (Fig. 3). Likewise, the three Salmonella Muenster isolates and the Salmonella 3,10:e,h:- isolate showed two similar patterns with only one band difference. The Salmonella 3,10:e,h:- isolate, recovered from M1 stage, showed an identical pattern to one of the Salmonella Muenster isolates, confirming its serovar. The XbaI-PFGE patterns obtained were different from those of other Salmonella Muenster isolates present in the Salmonella PFGE National Database. (Fig. 4).

FIG. 3.

FIG. 3.

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Dendrogram showing the genetic relationship between Salmonella Bovismorbificans isolated from pigs at different stages of production compared with Salmonella pulsed-field gel electrophoresis (PFGE) National Database.

FIG. 4.

FIG. 4.

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Dendrogram showing the genetic relation between Salmonella Muenster isolated from pigs at different stages of production compared with Salmonella PFGE National Database.

Discussion

Concerns about pork safety have stimulated research along the food chain in an attempt to determine the effects of various pre- and postharvest processes on the prevalence of Salmonella spp. contamination. Because of the temporal variability in prevalence within groups, longitudinal sampling is recommended for evaluating the behavior of Salmonella spp. in different farm systems (Funk et al., 2005).

In this surveillance, a low number of shedder sows at farrowing was observed. Additionally, stress factors are considered one of the triggers for Salmonella spp. shedding process; in this study, farrowing seems not to increase the shedding rate. Further, Funk et al. observed a decreased rate of shedding of Salmonella spp. between late gestation and early lactation. This reduction in shedding may be explained for certain changes in sow's environment such as diet, housing, water delivery, and husbandry practices (Hurd et al., 2003; Funk and Gebreyes, 2004; Nollet et al., 2005b). On the other hand, this low shedding rate could be related to the intermittent dissemination of the bacteria (Haye et al., 1981; Griffith et al., 2006) and cohort temporal variation (Kranker et al., 2003). Moreover, the seropositive sow's rate was 96%, which differs from that reported by Kranker et al. (2003) and Silva et al. (2006). Further, there is no information available regarding the influence of serological status of sows at farrowing avoiding the dissemination of the bacteria. Findings of this study suggest that immune response might have reduced the shedding rate. Further studies are needed to understand the influence of serological status of sows on the epidemiology of Salmonella spp. dissemination. The low shedding rate observed at the early lactation stages in this study as well as previous studies suggests that sows play a more important role in Salmonella spp. transmission to the newborns than is actually considered (Kranker et al., 2003).

There are different concerns regarding the bacteriological status in the lactation period. In this study, before weaning, 9% of piglets were shedding. This elimination rate is similar to the rate reported by Funk et al. (2001) for the same period (<7%). Other studies found no shedding piglets at weaning (Kranker et al., 2003; Silva et al., 2006). This information complements existing evidence, indicating that Salmonella spp. infection can occur in young pigs (Nollet et al., 2005a; Griffith et al., 2006). The percentage of seropositive piglets (48.8%) at 21 ± 3 days old may reflect the passive immune protection against Salmonella spp. infection (Davies et al., 1999). Moreover, this high percentage of seropositive piglets explains the moderate percentage of shedding piglets at weaning (Kranker et al., 2003).

At 35 ± 3 days old, midway through the postweaning period, 90% of the fecal samples were positive to Salmonella spp. This represents a high percentage compared with studies carried out in Denmark, where only 50% of piglets were positive using rectal swabs (Kranker et al., 2003). Rectal swab has less sensitivity than pooled floor fecal samples (Nollet et al., 2005b). Not only could the discrepancies observed be due to the sampling method (Kranker et al., 2003), but also due to the minor differences in the sensitivity of the bacteriological technique used. The high percentages of shedding pigs found in this surveillance might represent the real situation at weaning due to the all-in/all-out policies used. The rapid increase in Salmonella spp. prevalence in postweaning might have been triggered by weaning stress and associated factors, such as a change in feed and commingling of litters (Kranker et al., 2003; Griffith et al., 2006). On the other hand, horizontal transmission occurred at late lactation and early weaning increased the percentage of shedding pigs in postweaning period (Kranker et al., 2003).

One week before the pigs left the postweaning unit (65 ± 3 days old) the percentage of culture positive to Salmonella spp. fell drastically to 10%. This finding differs from the results of Kranker et al., 2003. This decrease in the number of shedding pigs is related to management practices. The effect of antibiotics on the frequency and length of shedding of Salmonella spp. has received little attention in pigs (Griffith et al., 2006). However, the common use of antibiotics as growth promoters or preventive of respiratory diseases could decrease the shedding rate of Salmonella spp.

Only 2.04% of the pigs were seropositive at 65 ± 3 days of age. The serological technique used in this study does not distinguish between maternal or pig's own antibodies. Thus, the fall of colostral antibodies could therefore mask an active immune response of pigs (Beloeil et al., 2003). On the other hand, the shedding rate observed at this point could not be enough to trigger an active immune response in a herd. The late nursery stage seems not to be important in the epidemiology of Salmonella spp. (Silva et al., 2006). Moreover, stress factors at the beginning of the nursery period were diminished at this point, and this could be the reason for the low shedding and serological rates observed. However, surveillance at this stage should be considered an important prior step in the early finishing period.

The bacteriological results obtained at the early finishing period showed a significant increase from the late nursery. The farm studied has its finishing units 5 km away from nursery facilities. Thus, relocation, among other stress factors, could be the reason for the increase in the shedding rate observed at 86 days of age (Davies et al., 1999). Further, the low number of positive pigs spreading the bacteria at the late nursery period seems to be the source of infection in early finishing units (Kranker et al., 2003). The Salmonella spp. shed-ding pattern observed in this study at the finishing stage suggests that the early finishing period plays a central role in the infection of finisher pigs. The dissemination pattern observed throughout this period agrees with that obtained by Funk et al. (2001) and Silva et al. (2006). However, we must emphasize that the failure to detect Salmonella spp. in middle and late finishing period did not indicate that the previously infected pigs continued harboring the organism (Nielsen et al., 1995).

The serological results obtained at the fattening period suggest an active immune response against the natural infection produced at the beginning of this period (Beloeil et al., 2003). The onset and serological peak has been described between 7 and 30 days postnatural infection (Cappuccio et al., 2006); it supports the delay observed against the infection and the serological response observed in this surveillance.

Although Salmonella Typhimurium had been isolated from a systemic infection in a previous study in this farm (Cappuccio et al., 2006), only Salmonella Muenster and Salmonella Bovismorbificans were identified, and both are considered to be of low pathogenicity to pigs (Griffith et al., 2006). The absence of Salmonella Typhimurium isolation could be due to the low prevalence of this serovar present during the study, as well as the natural competition and competence of multiple serovars circulating concurrently on the farm. The relatively common isolation of multiple serovars from a single swine source is presented in a study from Sweden that documented the incidence of up to seven of concurrent serovars in several farms from 1993 to 1997. Interestingly, neither Salmonella Muenster nor Salmonella Bovismorbificans were isolated (Boqvist et al., 2003). Moreover, there are few other reports of Salmonella Muenster or Bovismorbificans clinically affecting swine, and the majority of case reports are isolates from healthy animals and abattoir carcasses (Lázaro et al., 2004), hence the need to conduct surveillance of these serovars during the growth period (Barber et al., 2002; Hurd et al., 2002). Additionally, both serovars observed in this longitudinal study were not related with clinical disease in pigs, and their presence should be considered a potential contaminant of final products.

Barber (Holmberg et al., 1984) suggested that Salmonella spp. are transmitted readily between many ecological compartments within the swine production system. However, more precise genetic analyses of isolates would be needed to support this conclusion. The findings of this study appear to support this notion as was evidenced by PFGE, since for both Salmonella serovars identified, a single subtype was found throughout the different stages analyzed. Further, identical or very closely related Salmonella Muenster subtypes were iden-tified in the sow and its piglets. For the 3,10:e,h:- isolate, the serovar could not be established because the isolate did not express the second flagellar antigen, but according to the antigenic formula, it is highly probable that this isolate also belonged to the Muenster serovar. This observation was further confirmed with the PFGE results.

In this study, molecular analysis was important to determine that the same subtype was circulating and remained at all stages where Salmonella spp. were recovered.

Several studies have indicated an increase in MDR (Douris et al., 2008). Moreover, the extensive use of antibiotics in farms for both therapeutic and growth promotion purposes has been considered to be an important trigger for the emergence of antibiotic-resistant pathogens (Poppof et al., 1990; Threlfall et al., 2000), and their real impact on public health continues to be a matter of debate. Antibiotic-resistant strains have emerged worldwide and have led to increasing consumer concerns about animals for food consumption. Infections caused by resistant strains have shown a higher fatality rate compared to those caused by susceptible microorganisms (Threlfall et al., 2000). MDR were also more frequently associated with certain serovars (Zhao et al., 2007). In this study, three multidrug resistance strains (one belonged to Salmonella 3,10:e,h:- and two belonged to Salmonella Muenster) were identified. The MDR pattern appears only in the isolates belonging to sows and at 35 ± 3 days old. However, the resistance profile found is different from the one described in Brazil for the same serovar (Barber et al., 2002), which most likely reflects both different animal production environments and associated uses of antimicrobials in preventing or treating common diseases (Zhao et al., 2007).

Epidemiologic prevalence and behavior of Salmonella spp. infection can differ markedly between different farms and with different management practices. Importantly, this study allows us to determine the critical points of Salmonella spp. infection, their serological dynamics, and their shedding patterns. When all-in/all-out management production systems are utilized, residual infection of small numbers of animals within every group of raised pigs in the farrowing and nursery units can be considered a critical point in the persistence of subclinical Salmonella spp. infection. The association between stress and Salmonella spp. excretion has been previously described (Kranker et al., 2003; Nollet et al., 2005a; Griffith et al., 2006). Although there is no information about the degree of impact of transportation and commingling of litters, in three separate sites with all-in/all-out management systems, it was determined that transportation and litter commingling should be diminished to reduce Salmonella spp. excretion from the residual infected pigs. Moreover, further studies should be carried out to evaluate the real impact of other stressors, such as facilities, food usage, or antibiotic usage, which could be acting as a hazard point of Salmonella spp. subclinical persistence. Longitudinal periodical studies could allow determination of the optimal point in time in which to take specific control measures, such as feed and/or water acidification, and vaccination or antibiotic therapy, thereby reducing therapy failures, loss of food animals, production costs, or antibiotic resistance problems. Moreover, PFGE allowed determining the clonal relationships among different serovars throughout all the production stages. Antibiotic resistance profiles should be periodically included due to public health concerns.

Disclosure Statement

No competing financial interests exist.

References

  1. Asai T. Fujii S. Osumi T. Otagiri Y. Namimatsu T. Sato S. Isolation and serological survey of Salmonella in pigs in Japan. J Vet Med Sci. 2002;64:1011–1015. doi: 10.1292/jvms.64.1011. [DOI] [PubMed] [Google Scholar]
  2. Barber DA. Bahnson PB. Isaacson R. Jones CJ. Weigel RM. Distribution of Salmonella in swine production ecosystems. J Food Prot. 2002;65:1861–1868. doi: 10.4315/0362-028x-65.12.1861. [DOI] [PubMed] [Google Scholar]
  3. Beloeil PA. Chauvin C. Proux K. Rose N. Queguiner S. Eveno E. Houdayer C, et al. Longitudinal serological responses to Salmonella enterica of growing pigs in a subclinically infected herd. Prev Vet Med. 2003;60:207–226. doi: 10.1016/s0167-5877(03)00126-0. [DOI] [PubMed] [Google Scholar]
  4. Best EL. Lindstedt BA. Cook A. Clifton Hadley FA. Threlfall EJ. Liebana E. Multiple-locus variable-number tandem repeat analysis of Salmonella enterica subsp. enterica serovar Typhimurium: comparison of isolates from pigs, poultry and cases of human gastroenteritis. J Appl Microbiol. 2007;103:565–572. doi: 10.1111/j.1365-2672.2007.03278.x. [DOI] [PubMed] [Google Scholar]
  5. Boqvist S. Hansson I. Nord Bjerselius U. Hamilton C. Wahlstrom H. Noll B. Tysen E, et al. Salmonella isolated from animals and feed production in Sweden between 1993 and 1997. Acta Vet Scand. 2003;44:181–197. doi: 10.1186/1751-0147-44-181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cappuccio J. Quiroga M. Machuca M. Piñeyro P. Arauz S. Pinto M. Vigo G, et al. Porcine dermatitis and nephropathy syndrome as a cause of death, retard growth and increase isolation of Salmonella typhimurium in grower finisher phases in three Argentinean farms. In: Nielsen JP, editor; Jorsal SE, editor. Proceeding of the International Pig Veterinary Society. Copenhagen, Denmark: International Pig Veterinary Society; 2006. p. 95. [Google Scholar]
  7. Clinical and Laboratory Standards Institute. 15th Informational Supplement: M100-S15. Wayne, PA: 2005. Disk diffusion. Performance standards for antimicrobial susceptibility testing; pp. 52–57. [Google Scholar]
  8. Davies P. Funk J. Morrow W. Fecal shedding of Salmonella by a cohort of finishing pigs in North Carolina. J Swine Health Prod. 1999;7:213–234. [Google Scholar]
  9. Douris A. Fedorka-Cray PJ. Jackson CR. Characterization of Salmonella enterica serovar Agona slaughter isolates from the animal arm of the National Antimicrobial Resistance Monitoring System-Enteric Bacteria (NARMS): 1997 through 2003. Microb Drug Resist. 2008;14:55–63. doi: 10.1089/mdr.2008.0787. [DOI] [PubMed] [Google Scholar]
  10. Ekeroth L. Alban L. Feld N. Single versus double testing of meat-juice samples for Salmonella antibodies, in the Danish pig-herd surveillance programme. Prev Vet Med. 2003;60:155–165. doi: 10.1016/s0167-5877(03)00122-3. [DOI] [PubMed] [Google Scholar]
  11. Funk J. Gebreyes W. Risk factors associated with Salmonella prevalence on swine farms. J Swine Health Prod. 2004;12:246–251. [Google Scholar]
  12. Funk JA. Davies PR. Nichols MA. Longitudinal study of Salmonella enterica in growing pigs reared in multiple-site swine production systems. Vet Microbiol. 2001;83:45–60. doi: 10.1016/s0378-1135(01)00404-7. [DOI] [PubMed] [Google Scholar]
  13. Funk JA. Harris IT. Davies PR. Comparison of fecal culture and Danish Mix-ELISA for determination of Salmonella enterica subsp. enterica prevalence in growing swine. Vet Microbiol. 2005;107:115–126. doi: 10.1016/j.vetmic.2005.01.006. [DOI] [PubMed] [Google Scholar]
  14. Griffith R. Schawrtz K. Meyerholz D. Salmonella. In: Straw B, editor; Zimmerman J, editor; D'Allaire S, editor; Taylor D, editor. Diseases of Swine. Ames, Iowa: Blackwell Publishing Ltd; 2006. pp. 739–754. [Google Scholar]
  15. Ha Y. Jung K. Kim J. Choi C. Chae C. Outbreak of salmonellosis in pigs with postweaning multisystemic wasting syndrome. Vet Rec. 2005;156:583–584. doi: 10.1136/vr.156.18.583. [DOI] [PubMed] [Google Scholar]
  16. Haye SN. Kornegay ET. Mahan DC. Antibody response and serum protein and immunoglobulin concentrations in pigs from sows fed different protein sequences during gestation and lactation. J Anim Sci. 1981;53:1262–1268. doi: 10.2527/jas1981.5351262x. [DOI] [PubMed] [Google Scholar]
  17. Holmberg SD. Osterholm MT. Senger KA. Cohen ML. Drug-resistant Salmonella from animals fed antimicrobials. N Engl J Med. 1984;311:617–622. doi: 10.1056/NEJM198409063111001. [DOI] [PubMed] [Google Scholar]
  18. Hunter SB. Vauterin P. Lambert-Fair MA. Van Duyne MS. Kubota K. Graves L. Wrigley D, et al. Establishment of a universal size standard strain for use with the PulseNet standardized pulsed-field gel electrophoresis protocols: converting the national databases to the new size standard. J Clin Microbiol. 2005;43:1045–1050. doi: 10.1128/JCM.43.3.1045-1050.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hurd H. Mcken J. Griffith R. Rostagno M. Estimation of the Salmonella enterica prevalence in finishing swine. Epidemiol Infect. 2003;132:127–135. doi: 10.1017/s0950268803001249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hurd HS. McKean JD. Griffith RW. Wesley IV. Rostagno MH. Salmonella enterica infections in market swine with and without transport and holding. Appl Environ Microbiol. 2002;68:2376–2381. doi: 10.1128/AEM.68.5.2376-2381.2002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Koneman E. Allen S. Janda W. Schreckenberger P. Winn W. Diagnóstico Microbiológico. 5th. Buenos Aires, BA: Editorial Médica Panamericana S.A. Springer; 1999. (In Spanish.) [Google Scholar]
  22. Kranker S. Alban L. Boes J. Dahl J. Longitudinal study of Salmonella enterica serotype Typhimurium infection in three Danish farrow-to-finish swine herds. J Clin Microbiol. 2003;41:2282–2288. doi: 10.1128/JCM.41.6.2282-2288.2003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Lanza I. Control of infectious enteric diseases of swine. In: Done S, editor; Thomson J, editor; Varley M, editor. Proceeding of the 15th International Pigs Veterinary Society. Birmingham, England: International Pig Veterinary Society; 1998. pp. 79–85. [Google Scholar]
  24. Lázaro N. Tibana A. Reis E. Rodrigues D. Quintaes B. Hofer E. Padrão de susceptibilidade a anitimicriobianos e perfil plasmidial em Salmonella Muenster isoladas de suínos e do ambiente de abatadeuros. Pesqui Vet Bras. 2004;24:65–70. (In Portuguese.) [Google Scholar]
  25. Liebana E. Molecular tools for epidemiological investigations of S. enterica subspecies enterica infections. Res Vet Sci. 2002;72:169–175. doi: 10.1053/rvsc.2001.0550. [DOI] [PubMed] [Google Scholar]
  26. Lo Fo Wong DM. Dahl J. van der Wolf PJ. Wingstrand A. Leontides L. von Altrock A. Recovery of Salmonella enterica from seropositive finishing pig herds. Vet Microbiol. 2003;97:201–214. doi: 10.1016/j.vetmic.2003.09.012. [DOI] [PubMed] [Google Scholar]
  27. Nielsen B. Baggesen D. Bager F. Haugegaard J. Lind P. The serological response to Salmonella serovars typhimurium and infantis in experimentally infected pigs. The time course followed with an indirect anti-LPS ELISA and bacteriological examinations. Vet Microbiol. 1995;47:205–218. doi: 10.1016/0378-1135(95)00113-1. [DOI] [PubMed] [Google Scholar]
  28. Nollet N. Houf K. Dewulf J. De Kruif A. De Zutter L. Maes D. Salmonella in sows: a longitudinal study in farrow-to-finish pig herds. Vet Res. 2005a;36:645–656. doi: 10.1051/vetres:2005022. [DOI] [PubMed] [Google Scholar]
  29. Nollet N. Maes D. Duchateau L. Hautekiet V. Houf K. Van Hoof J. De Zuttera L, et al. Discrepancies between the isolation of Salmonella from mesenteric lymph nodes and the results of serological screening in slaughter pigs. Vet Res. 2005b;36:545–555. doi: 10.1051/vetres:2005014. [DOI] [PubMed] [Google Scholar]
  30. Oliveira CJ. Carvalho LF. Fernandes SA. Tavechio AT. Domingues FJ., Jr Prevalence of pigs infected by Salmonella Typhimurium at slaughter after an enterocolitis outbreak. Int J Food Microbiol. 2005;105:267–271. doi: 10.1016/j.ijfoodmicro.2005.04.016. [DOI] [PubMed] [Google Scholar]
  31. Poppof M. Bockemuhl J. McWorther-Murlin A. Res Microbiol. Unité des Entérobacteries, U199 INSERM. Paris: Institut Pasteur; 1991. Supplement 1990 (n°34) to the Kauffman-White scheme; pp. 1029–1033. [Google Scholar]
  32. Ribot EM. Fair MA. Gautom R. Cameron DN. Hunter SB. Swaminathan B. Barrett TJ. Standardization of pulsed-field gel electrophoresis protocols for the subtyping of Escherichia coli O157:H7, Salmonella, and Shigella for PulseNet. Foodborne Pathog Dis. 2006;3:59–67. doi: 10.1089/fpd.2006.3.59. [DOI] [PubMed] [Google Scholar]
  33. Ridley AM. Threlfall EJ. Rowe B. Genotypic characterization of Salmonella enteritidis phage types by plasmid analysis, ribotyping, and pulsed-field gel electrophoresis. J Clin Microbiol. 1998;36:2314–2321. doi: 10.1128/jcm.36.8.2314-2321.1998. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sauli I. Danuser J. Geeraerd AH. Van Impe JF. Rufenacht J. Bissig-Choisat B. Wenk C, et al. Estimating the probability and level of contamination with Salmonella of feed for finishing pigs produced in Switzerland—the impact of the production pathway. Int J Food Microbiol. 2005;100:289–310. doi: 10.1016/j.ijfoodmicro.2004.10.026. [DOI] [PubMed] [Google Scholar]
  35. Schawrtz K. Proceeding of the 28th American Association of Swine Practitioners. Quebec, Canada: American Association of Swine Practitioners; 1997. Salmonella: what are the best approaches? pp. 385–388. [Google Scholar]
  36. Silva L. Gotardi C. Vizzotto R. Kich J. Cardoso M. Infecção por Salmonella enterica em suínos criados em um sistema integrado de produção do sul do Brasil. Arq Bras Med Vet Zootec. 2006;58:455–461. (In Portuguese.) [Google Scholar]
  37. Threlfall EJ. Ward LR. Frost JA. Willshaw GA. The emergence and spread of antibiotic resistance in food-borne bacteria. Int J Food Microbiol. 2000;62:1–5. doi: 10.1016/s0168-1605(00)00351-2. [DOI] [PubMed] [Google Scholar]
  38. Vigo G. Cappuccio J. Piñeyro P. Machuca M. Quiroga M. Moredo F. Giacoboni G, et al. Proceeding of the 13th Congreso Brasileño de Veterinarios Suinos. Florianópolis; Brasil: 2007. Bacteriological and serological longitudinal study of Salmonella enterica in a positive three site farrow to finish farm; pp. 16–19. [Google Scholar]
  39. Vigo G. Leotta G. Caffer M. Sanguinetti H. Piñeyro P. Cappuccio J. Perfumo C. Proceeding of the 2nd Congreso Latinoamericano de Suinicultura. Foz do Iguacu; Brasil: 2004. An outbreak of Salmonella Typhimurium in pigs in a farm with persistent PCV-2 associted diseases; p. 505. [Google Scholar]
  40. Weiss L. Nonig R. Cardoso M. da Costa M. Occorrencia de Salmonella spp. em suínos de terminação no Rio Grande do Sul. Pesq Vet Bras. 2002;22:104–108. (In Portuguese.) [Google Scholar]
  41. Zhao S. McDermott PF. White DG. Qaiyumi S. Friedman SL. Abbott JW. Glenn A, et al. Characterization of multidrug resistant Salmonella recovered from diseased animals. Vet Microbiol. 2007;123:122–132. doi: 10.1016/j.vetmic.2007.03.001. [DOI] [PubMed] [Google Scholar]

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