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African Journal of Pig Farming ISSN 2375-0731 Vol. 5 (8), pp. 001-007, August, 2017. Available online at 
www.internationalscholarsjournals.org © International Scholars Journals 

 

Author(s) retain the copyright of this article. 

 

Full Length Research Paper 

 

Antibiotic resistance, integrons and Salmonella 

genomic island 1 among Salmonella Schwarzengrund 

in broiler chicken and pig 
 

Yu-Chih Wang1, Yi-Chih Chang1,6, Hsiao-Li Chuang2, Chien-Chao Chiu2, Kuang-Sheng Yeh4, 
Chao-Chin Chang3, Shih-Ling Hsuan5 and Ter-Hsin Chen3,5*  

1
Department of Veterinary Medicine, National Chung Hsing University, Taichung, 402, Taiwan. 

2
National Laboratory Animal Center, National Applied Research Laboratories, Taipei, 115, Taiwan.  

3
The Graduate Institute of Microbiology and Public Health, College of Veterinary Medicine, National Chung 

Hsing University, Taichung, 402, Taiwan.  
4
Department of Microbiology and Immunology, School of Medicine, Taipei Medical University, Taipei, 402, 

Taiwan. 
5
Graduate Institute of Veterinary Pathobiology, College of Veterinary Medicine, National Chung 

Hsing University, Taichung, 402, Taiwan.  
6
Department of Medical Laboratory Science and Biotechnology, China Medical University, Taichung, 402, Taiwan. 

 
Accepted 04 March, 2017 

 
Salmonella Schwarzengrund is an infrequent serovar isolated from humans in Taiwan. However, this serovar 
is which is highly invasive, is found in humans as report by two independent epidemiologic surveys. The 
present study delineated the widespread resistance to fluoroquinolone in S. Schwarzengrund isolated from 
both poultry and swine carcesses in Taiwan. We conducted the present study to investigate the prevalence 
and characteristics of S. Schwarzengrund isolated from broiler chicken in central Taiwan. A total of 187 S. 
Schwarzengrund isolates was gotten from slaughtered chicken and pigs in central Taiwan between June 
2006 and March 2007. The percentages of resistance were as follows: ampicillin (90.74%), chloramphenicol 
(69.75%), florfenicol (24.07%), streptomycin (91.98%), trimethoprim and sulfamethoxazole (91.36%), nalidixic 
acid (96.30%), ciprofloxacin (8.02%), tetracycline (95.68%), amikacin (8.64%), ceftiofur (0.62%) and ceftriaxone 
(0%). Four types of class 1 integrons were detected: 1.2 kb carried blaPSE-1, aadA2 gene (n = 2), 1.2 kb 
carried blaPSE1, dfraA1 gene (n = 2), 1.0/1.2 kb carried blaPSE-1, aadA2 ( n = 2) and 1.9 kb carried dfrA12, 
aadA2 gene (n = 152). XbaI-digested PFGE patterns generated related clusters implicated in the 
dissemination of integron. Thirteen ciprofloxacin-resistant isolates were all detected in 5 identical mutations 
in the QRDR of gyrA, parC and parE. Our results suggest that there is a risk of transmitting multidrug 
resistant Salmonella between pig and chicken farms, and also suggest that there is a need to prevent the 
transmission of this organism from a neighboring contaminated farm. 

 

Key words: Integron, multidrug resistance, Salmonella Schwarzengrund. 
 
 
INTRODUCTION 
 
Salmonella enterica is one of the most common causes 

of human foodborne infection. Most serovars of Salmo-

nella infections result to self-limited gastroenteritis which 

is usually recovered from without treatment. However  
 
 
 
*Corresponding author. E-mail: thc@dragon.nchu.edu.tw. Tel: 
886 4 22840894. Ext: 407. 

 
 
 
 

 
some lead to severe invasive infections. Invasive 
Salmonella infections can be life threatening and usually 

require hospitalization and antibiotics treatment. 
Antimicrobial- resistant Salmonella were associated with 

an increased rate of hospitalization compared with out-
breaks caused by multidrug resistant (MDR) Salmonella 
strains (Boyd et al., 2002) . In Taiwan, Salmonella 

Schwarzengrund was the 3rd or 4th frequent serotypes 
(Chiu et al., 1999; Lauderdale et al., 2006), and the high 



 
 
 

 

prevalence of resistance to extended-spectrum 
cephalosporins or fluoroquinolones found in Salmonella 
Schwarzengrund (Chen et al., 2006; Lauderdale et al., 
2006; Yan et al., 2005) has become an important 
concern. This serovar showed highly invasive rate (25 
and 12.5%, respectively, for) report by two independent 
epidemiologic surveys (Lauderdale et al., 2006; Vugia et 
al., 2004).  

Multidrug resistance of Salmonella is linked to the 
presence of class 1 integrons (Leverstein et al., 2003). 
The acquisition of integrons via horizontal gene transfer 
allows bacteria to rapidly evolve. Class 1 integrons are 
the most common integrons found in clinical Salmonella 
isolates (Fluit 2005; Kwon et al., 2002) . Salmonella 
genomic island 1 (SGI1) is a 43 kb genomic island, which 
contains a complex integron. SGI1 is mobile and can be  
transferred between di erent Salmonella serovars and 

other bacteria in the presence of a helper plasmid 
(Doublet et al., 2005; Khemtong and Chuanchuen, 2008).  

Studies suggest that broiler chickens are a major 
reservoir for Salmonella Schwarzengrund (Aarestrup et 
al., 2007; Bangtrakulnonth et al., 2004). However, 
epidemiology data of MDR Salmonella Schwarzengrund 
among chickens are limited. One potential health hazard 
associated with the veterinary use of antibiotics is the 
transmission of antibiotic-resistant pathogens from 
animals to humans, this study examined antibiotic 
resistance of Salmonella Schwarzengrund isolated from 

broiler and swine in Taiwan. 
 

 
MATERIALS AND METHODS 
 
Sample isolates 
 
A total of 187 Salmonella Schwarzengrund isolates were obtained 
from slaughtered chickens from 5 chicken slaughterhouses (N = 
159) in 2006 - 2007 and 2 pigs slaughterhouses (N = 28) in 2005 - 
2006 in central Taiwan. Isolates were serotyped by using 
commercial Salmonella O and H antisera purchased from S&A 
Reagents Laboratory (Bangkok, Thailand) and Denka Seiken 
(Tokyo, Japan). All isolates were stored in 20% glycerol at -80°C. 
Bacteria were grown on Tryptic Soy agar/broth (Difco, MI, U.S.A.) at 
37°C. 

 

Antibiotic susceptibility test 
 
Antimicrobial susceptibility was tested by a standard disk diffusion 
method, and Escherichia coli ATCC 25922 was used for control. 
The antimicrobial agents used were ampicillin, chloramphenicol, 
florfenicol, streptomycin, trimethoprim-sulfamethoxazole, nalidixic 
acid, ciprofloxacin, amikacin, ceftiofur, ceftriaxone and tetracycline. 
Susceptible and resistant isolates were defined according to the 
criteria suggested by the Clinical and Laboratory Standards 
Institute. 

 

Genotyping by pulsed-field gel electrophoresis (PFGE) 
 
Genotypes of all Salmonella Schwarzengrund isolates were 

determined by pulsed-field gel electrophoresis (PFGE) analysis 

 
 
 
 

 
using the restriction endonuclease XbaI to digest total genomic 
DNA. The procedure for the PFGE was performed according to the 
standard protocol developed by the Centers for Disease Control 
and Prevention (Barrett et al., 2006). The digested DNA was 
separated by the use of CHEF DR II (Bio-Rad) in 0.5x Tris-borate-
EDTA at 14°C for 20 h. 
 
 
PCR analysis, DNA purification and DNA sequencing 
 
DNA templates used for PCR were prepared by boiling bacterial 
cultures or by using the QIAGEN Genomic-tip System (QIAGEN). 
Amplifications were performed in 25 L reaction mixtures containing 
2.5 L of DNA, 2.5 L 10X PCR buffer, 1.5 M MgCl2, 200 M each 
deoxynucleoside triphosphate, 2.5 U of Taq DNA Polymerase 
(Promega) and 1 M each primer. To amplify fragments larger than 3 
kb, Blend Taq-Plus polymerase (TOYOBO) was used instead of 
Taq DNA polymerase. The PCR products were visualized by 
ethidium bromide staining after agarose gel electrophoresis. 
Amplification products were purified with the QIAquick PCR 
Purification Kit (Qiagen) for sequencing. The resulting DNA 
sequence data were compared to the GenBank Database using the 
Blast algorithm available at the National Center for Biotechnology 
for Biotechnology information website (www.ncbi.nlm.nih.gov).  

Class 1 integrons were screened by PCR with primers 5’ -CS and 
3’ -CS as described previously (Lévesque et al., 1995). Detection of 
Salmonella genomic island 1 was done using primers 
corresponding to left and right junctions as described previously 
(Cloeckaert et al., 2000). PCR mapping of the typical antibiotic 
resistance genes associated with SGI1 was performed using 
conditions and primers described previously (Boyd et al., 2002; 
Levings et al., 2005).  

For the detection of resistance-mediating mutations in the DNA 
gyrase, topoisomerases IV and plasmid- mediated quinolone 
resistance genes, the quinolone-resistance determining region 
(QRDR) of gyrA, gyrB, parC, parE, qnrA, qnrB and qnrS genes was 
amplified by PCR oligonucleotide primers which were amplified and 
sequenced. The sequences of the primers and PCR condition was 
described previously (Casin et al., 2003; Gay et al., 2006; Ling et 
al., 2003). 
 

 

RESULTS 

 

Antimicrobial susceptibility testing 

 

All isolates were sensitive to ceftriaxone and only one 
isolate was resistant to ceftiofur. Eighty-six isolates 
(45.99%) displayed the pentaresistance phenotype 
(ACSSuT). Resistance to nalidixic acid, tetracycline,  
streptomycin, trimethoprim- sulfamethoxazole and 
ampicillin was found in 96.30, 95.68, 91.98, 91.36 and 
90.74% of the isolates, respectively. Isolates from 
chickens resistant to ampicillin, trimethoprim-
sulfamethoxazole, nalidixic acid and tetracycline were 
more than the isolates from pigs. However, resistance to 
florfenicol, ciprofloxacin and amikacin of the isolates from 
pigs was higher than isolates from chickens (Table 1). 
 

 

PCR detection of integrons and gene cassettes Class 

1 integrons were positive in 84.49% (158/187) 



       
 

Table 1. Percentages of Salmonella Schwarzengrund isolates resistant to antibiotics from broiler chickens and pigs.  
 

        
 

 
Antibiotic 

  % of resistant isolates from    
 

 

Broiler(n = 159) Pig (n = 28) Overall (n = 187) 
 

 

   
 

 Ampicillin 93.06 (134) 72.22 (13) 90.74   
 

 Chloramphenicol 69.44 (100) 72.22 (13) 69.75   
 

 Florfenicol 18.75 (27) 66.67 (12) 24.07   
 

 Streptomycin 91.67 (132) 94.44 (17) 91.98   
 

 Trimethoprim-Sulfamethoxazole 94.44 (136) 66.67 (12) 91.36   
 

 Nalidixic acid 97.92 (141) 83.33 (15) 96.30   
 

 Ciprofloxacin 4.17 (6) 38.89 (7) 8.02   
 

 Tetracycline 96.53 (139) 88.89 (16) 95.68   
 

 Amikacin 1.39 (2) 66.67 (12) 8.64   
 

 Ceftiofur 0.69 (1) 0 0.62   
 

 Ceftriaxone 0  0 0   
 

 
 

 
Table 2. Characteristics of the serovar Schwarzengrund strains tested in this study.  

 
 Size(kb)  of  integron  I  and 

Variant genomic island name 
Isolated from 

No. of strains  

 

identity of resistance gene Broiler pig 
 

   
 

 None - 10 4 10 
 

 1.2- blaPSE-1, aadA2 SGI1 1 1 2 
 

 1.2- blaPSE1, dfraA1 SGI1-F 1 1 2 
 

 1.2- blaPSE-1, 1.0- aadA2 SGI1 2 0 2 
 

 1.9- dfrA12, aadA2 - 130 22 152 
 

 
 

 

isolates with 3 PCR amplification product patterns: 1,900 

(152 isolates), 1,200 (4 isolates) and 1,000 + 1,200 bp (2 

isolates) detected. Sequence analysis of the integron PCR 

products showed the presence of classic gene cassettes in 

the integrons, including aadA2 (which confer resistance to 

streptomycin), the dihydrofolate reductase gene cassettes 

dfrXII, dfrA1 (which confer resistance to trimethoprim) and 

the beta-lactamase gene blaPSE1 (which confers resistance 

to ampicillin) (Table 2). Six isolates were positive to the 

SGI1. PCR mapping of the antibiotic resistance gene cluster 

demonstrated that 4 isolate contain the SGI1 and 2 isolate 

contain SGI1-F (Table 2). 

 
 

 

chicken and pig sources. 
For all isolates of Salmonella Schwarzengrund, 

screening gave negative results for qnrA, qnrB and qnrS 
genes. And 13 ciprofloxacin-resistant isolates were all 
detected as double mutations in the QRDR of gyrA (S83F 
and D87G), double mutations in the QRDR of parC (T57S 

and S80R) and an additional mutation in the QRDR of 
parE (S458P). These 13 ciprofloxacin-resistant 
Salmonella Schwarzengrund isolates were isolated from 
2 different chicken slaughterhouses and 2 different pig 
slaughterhouses. All these isolates had the same 
genotype as 2D. 
 

 

Genotype of PFGE 

 

Eleven PFGE profiles were observed from the 187 
isolates in the study when total genomic DNA was 
digested with XbaI. Genotype 2D was seen in 84 isolates 

(44.92%) and genotype 2B was represented by 44 
isolates (22.53%) . The third common genotype was 1A 
which was found in 15 isolates (8.02%). A dendrogram 
generated from Dice coefficients of similarity indicated 
that all isolates were highly related with Dice values 
greater than 84% (Figure 1). The predominant genotypes 
(1A, 2B and 2D) were observed in isolates from both 

 
 
DISCUSSION 

 

Nontyphoidal Salmonella which causes foodborne 

diseases has become an important public health problem 
worldwide. Salmonella Enteritidis and Salmonella 
Typhimurium were the most common causes of non-
typhoidal salmonellosis. Salmonella Schwarzengrund is a 
less common cause of human salmonellosis worldwide. 
However, the incidence of Salmonella Schwarzengrund 

has increased in recent years (Bangtrakulnonth et al., 
2004; Boyd et al., 2002). This serovar is commonly found 
from invasive salmonellosis patients in Taiwan 



  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
Figure 1. PFGE patterns of XbaI-digested chromosomal DNA of S. enterica serovar Schwarzengrund. The dendrogram was 

constructed by use of the UPGMA algorithm and the Dice similarity coefficient by using Gelcompare II software with 1% 

position tolerance. 

 

(Boyd et al., 2002). 
This study showed a high frequency of antimicrobial 

drug resistance, including ampicillin, trimethoprim-
sulfamethoxazole, among Salmonella Schwarzengrund 
isolates from slaughtered chickens. In contrast, the 
frequency of resistance to florfenicol, ciprofloxacin and 
amikacin of isolates from pigs was higher. It may 
correlate the antibiotics used in the pig farms and broiler  
farms. 13 ciprofloxacin-resistant Salmonella 
Schwarzengrund isolates found in 2 different chicken 
slaughterhouses and 2 different pig slaughterhouses had 
the same mutations in the QRDRs of gyrA, par and 
identical PFGE pattern. Molecular characterization 
suggests an epidemiological relationship between the 
swine and chicken Salmonella Schwarzengrund isolates. 
Our results suggest that there is a risk of transmitting 
multidrug resistant Salmonella between pig and chicken 
farms, and also suggest that there is a need to prevent 
the transmission of this organism from a neighboring 
contaminated farm.  

This study also found the widespread occurrence of class 

1 integrons and SGI1 variants in multidrug resistant 

Salmonella Schwarzengrund isolates from broiler chickens 

and pigs. Genes encoding for resistance to trimethoprim 

(dfrA12) and aminoglycosides (aadA2) were most commonly 

found. The presence of this 1.9 kb integrons has been found 

in many serovars worldwide (Doublet et al., 2005; Fluit 2005; 

Gay et al., 2006; Hsu et al., 2006). The transfer of 

conjugative plasmids is a common mechanism for genetic 

exchange between bacteria, transfer can occur both within 

bacterial species and between different species. Therefore, 

the presence of the integron-carrying Salmonella isolates 

detected in this study could have contributed not 

 

 

only to the spread of drug-resistant strains but also to the 

spread of mobile elements carrying drug resistance 

genes across bacterial species. 
 

 
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