Research Article: Salmonella Typhimurium in Iran: Contribution of molecular and IS200 PCR methods in variants detection

Date Published: March 13, 2019

Publisher: Public Library of Science

Author(s): Reza Farahani Khaltabadi, Nader Shahrokhi, Mina Ebrahimi-Rad, Parastoo Ehsani, Joël Mossong.


Salmonella Typhimurium, a zoonotic pathogen, is regarded as a major health and economic concern worldwide. Recently, monophasic variants of this serovar have been significantly associated with human gastroenteritis outbreaks globally, making its accurate identification essential for epidemiological and control purposes. We have identified and analyzed 150 S. Typhimurium from 884 Salmonella genus isolated from humans, domestic animals, poultry, food items and abattoirs origins. The Salmonella isolates were obtained from Iranian National Veterinary Reference Laboratories of 9 provinces during 2007–2016, and from five hospitals in Tehran in 2015. The isolates were evaluated biochemically, serologically, and by PCR amplification of invA, mdh, STM4492, fliC, fljA, fljB, hin genes, IS200 and DT104. invA and mdh genes were used to confirm the S. Typhimurium serotype, fliC and fljB genes for determination of monophasic variants and amplification of IS200 to discriminate the monophasic variants from the closely related serotypes. We identified 78.6% (118/150) as classical S. Typhimurium (fliC, fljB and IS200 positive), 12.6% (19/150) were IS200 negative from all isolates. DT104 is another marker for S.Typhimurium serovar typing. Contrary to EFSA guidelines 20.6% (19/29) of human isolates that lacked IS200 insertion sequence, were confirmed as S.Typhimurium. Compared to the North American/European isolates the low prevalence of fljB negative 6% (9/150) and the high abundance of fliC negative 23.3% (35/150) isolates also were indicative of a different regional atypical population. Studies have shown that the prevalence of monophasic (fljB-) S. Typhimurium worldwide is promoted by the Swine industry. Thus, one reason for this high number of different atypical strains could be inhibition of swine breeding system (house hold and industry) in Iran. These results demonstrate a need for a modified identifying protocol to overcome the regional differences.

Partial Text

Non-typhoidal Salmonellae are a common cause of salmonellosis in humans and animals. Despite improvements in hygiene and food safety standards, the infection remains a common public health concern worldwide [1]. The spreading of infectious human Salmonella serovars occurs through consumption of contaminated food and water, contact with infected animals, international food trades and travelling by infected people within and among countries [2]. The burden of the infection is not limited to the associated morbidity and mortality costs of humans and animals, but it also includes the loss of trade and the consequent socioeconomic problems [2, 3]. Salmonella, enterica and bongori, are the two species of the genus with the former containing six subspecies enterica, salamae, arizonae, diarizonae, houtenae and indica. The most clinically important is S.enterica sub species enterica due to its common association with humans and warm-blooded animals and includes approximately 1500 serovars [4, 5]. Although potentially, all serovars of this subspecies could be pathogenic for humans, only about 80 of them account for nearly 99% of Salmonella infections in humans and domestic animals. Salmonella Typhimurium ranks among the top-five serovars, isolated from pigs, poultry, red meat and food-borne outbreaks in the European Union and the United States [4–8]. In Iran, the latest available data by the Center for Communicable Diseases and Control in 2011 showed that from 1038 reported food-borne diseases outbreaks, 5.1% were caused by Salmonella isolates, all serovars taken together [9].

Out of 819 animals and 65 human Salmonellas isolates, 121 and 29 were positive for O4 (group B) antigen and PCR positive for invA and mdh genes respectively and were identified as S.Typhimurium (Table 1). The STM4492gene was not detected in 6.6% (10/150) of the strains. One of the STM4492 negative strains belonged to human isolates (Table 3), three to red meat (Table 4), one to sheep (Table 5), four to chicken and one to chicken paste isolates (Table 6). Sero-agglutination tests for phase 1 and 2 flagellar antigens were repeated on these isolates, confirming the original identification and phase 2 serotyping was positive for one human, one red meat and one sheep sample.

Effective surveillance and control of pathogens require accurate identification and strain characterization. In this study, 150 out of 884 Salmonella isolates were identified as S.Typhimurium based on conventional and PCR methods, of which 121 were of non-human origin sent to the National Veterinary Laboratory and 29 from human diarrheal cases.

Our results showed that S. Typhimurium with IS200deletion is a fairly common serotype among the Iranian Salmonella isolates of human and non-human origins. Therefore, with the procedure recommended by EFSA some of the variants will not be correctly identified and therefore missed. In addition, with 34.3% prevalence of DT104 subtype that could carry resistant genes, among the atypical strains highlight the importance of adaptation of new guidelines for identification of all the variants.




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