Isolation and Detection of Cronobacter sakazakii from Infant Dried Milk using PCR and RT-PCR techniques

Abstract

The present study is an attempt for detection of Cronobacter sakazakii by conventional PCR methods using species-specific primers and to detect the gene expression of thermotolerance gene (Tt gene) as virulence factor. 105 samples of  powdered infant formula (PIF)  and infant's food were collected  from  Baghdad Governorate and Mesan Governorate  markets from November  2016 to June 2017. The samples  were cultured on the selective media including Enterobacter sakazakii Isolation Agar(ESIA)  and trypton soy agar(TSA). Sixty eight  (64.76%) isolates appeared with  yellow pigment colonies when cultured on TSA and gave bluish - green colonies on chromogenic media (ESIA). After the growth of bacteria, isolates were identified by microscopic examination and biochemical tests. The identification of C. sakazakii confirmed that twenty one isolates identified as C. sakazakii and the others identified as Enterobacter spp. by using systems API 20E and VITEK2 systems. In addition, a molecular identification has been done by PCR technology utilizing 16S Rrna gene. The results showed that 42(61.76%) from 68 isolates were identified as C. sakazakii and 26(38.24%) were Enterobacter spp. The isolated  Enterobacter spp include  E. cloacae ssp. Cloacae, E. cloacae ssp. dissolvens, E. hormaechei and  E . kobei, E. ludwigii .