In silico analysis of genome sequences of control and well documented phage types of S. Typhimurium revealed two CRISPR loci, CRISPR-1 and CRISPR-2, within all phage types of S. Typhimurium. Although DRs are almost identical among all phage types of S. Typhimurium spacers sequences within the CRISPR loci are not unique to the phage type as strains belong to the same phage type have different spacers and subsequently different CRISPR/CRISPOL type (Table 1) furthermore, different phage types have identical spacers and same CRISPR/CRISPOL type (Table 2).
Different CRISPR/CRISPOL type within the same phage type of S. Typhimurium
In Table 1, three strains of S. Typhimurium that belong to phage type DT1 including strains DT1, TM 68-619 and TM 65-111 have different spacers and subsequently show different CRISPR/CRISPOL type; 8579/430, 2536/54 and 7387/90 respectively. Two strains belong to phage type DT10 have different CRISPR/CRISPOL type; MS34 (9509/1629) and S81-784 (9913/1688). Two strains belong to phage type DT15a have different CRISPR/CRISPOL type; 9517/1634 in isolate MS41 and 9916/1756 in isolate S81-798. Moreover, three strains belong to DT41 have different CRISPR/CRISPOL type; 9513/1630 in isolate M11-2004, 7434/223 in isolate CQ 41 and 9929/1766 in isolate S02-0321.
Identical CRISPR/CRISPOL type within different phage types of S. Typhimurium
CRISPR/CRISPOL type among phage types DT8 and DT30
Identical spacers were detected among different phage types of S. Typhimurium. For example, three strains of DT8 including M12-2001, M15-2006 and MS32 have the same CRISPR/CRISPOL type (812/250) as a strain belongs to phage type DT30 (MS57). Moreover, different strains belong to phage type DT8 have different CRISPR/CRISPOL type; M18-2003 (1069/6) and MS150057 (2260/708) (Table 2).
Interestingly, S. Typhimurium DT8 strains associated with the foodborne outbreak in the summer of 2013 in the States of Jersey [10] showed identical CRISPR/CRISPOL type (1069/6) however, the same CRISPR/CRISPOL type were reported in other DT8 strains that do not belong to the outbreak as confirmed by WGS [10]. Identical spacers were detected among outbreak associated and non-outbreak associated DT8 strains (Additional file 1: Table S1).
CRISPR/CRISPOL type among phage types DT104, DT104b and U302
Variations in the CRISPR/CRISPOL type among strains of the same phage type such as DT104 and DT104b have been also noticed (Table 2). Although three strains of S. Typhimurium phage type DT104 including TM75-339, MS150098 and MS150095, have identical spacer sequences and CRISPR/CRISPOL type (12/21) the same CRISPR/CRISPOL type is present in different phage types including U302 (M18-2006; 12/21) and DT104b (MS130531; 12/21).
CRISPR/CRISPOL type among phage types DT40, DT56, DT99 and U319
Strains of S. Typhimurium belong to different phage types such as DT99, DT56, U319 and DT40 (S05-2864) have identical spacer sequences and identical CRISPR/CRISPOL type (7433/14). Moreover, several strains belong to phage type DT40 including S05-2864, M20-2006, M19-2003 and CQ 40 have different CRISPR/CRISPOL type; 7433/14, 9520/1637, 9519/1636 and 745/18 respectively (Table 2).
CRISPR/CRISPOL type among phage types DT7a, DT20a, DT120, DT193 and untypable strains
In Table 2, strains of S. Typhimurium belong to phage type DT120 have different spacers and subsequently different CRISPR/CRISPOL type including S02-3776 (9921/1759), 07_2198 (9911/1753), M16-2000 (9510/1428), and S/20160374 (322/1).
Interestingly, a strains of phage type DT120 (M16-2000) has identical spacers and CRISPR/CRISPOL type (9510/1428) as another strain belongs to phage type DT7a (MS120840). Moreover, some strains belong to phage types DT120 (S/20160374 and S/20160407), DT20a (MS150110), DT193 (MS150007) and untypable strain (MS150097) have identical spacers and therefore share the same CRISPR/CRISPOL type (322/1). Different strains belong to phage type DT193 have different spacers and CRISPR/CRISPOL type; MS150007 (322/1) and MS150252 (317/2).
CRISPR/CRISPOL type among phage types DT3, DT12 and DT193a
Some strains of phage types DT12 (DT12) and DT3 (S81-482) have identical spacers and identical CRISPR/CRISPOL type; 5268/19. Moreover, a strain belongs to DT12 (S02-2651) has identical CRISPR/CRISPOL type, 774/46, as a strain belongs to phage type DT193a (MS120454) (Table 2).
CRISPR/CRISPOL type among phage types DT135, DT191a and RDNC
Identical spacer sequences and CRISPR/CRISPOL type (91/4) were detected in different phage types of S. Typhimurium including DT135 (MS150112 and MS150180), DT191a (DT19a) and strains that react with phages but do not confirm to recognized pattern (RDNC) (MS150102 and MS150230). Furthermore, other strains belong to phage type DT135 show different spacers and subsequently different CRISPR/CRISPOL type; 5753/396 in DT135 and 3247/66 in MS150100 (Table 2).
CRISPOL assay confirms the no relation among phage type and CRISPRs
CRISPOL assay developed by Fabre et al. [5] when carried out on representative phage types of S. Typhimurium it reveals that there is no relation among the phage type and the CRISPOL type as strains belong to the same phage type have different CRISPOL type as seen in DT104 strains (Table 3). On the other hand, different phage types including DT7, DT193, U311, DT41 showed identical CRISPOL type as ‘1’ (Table 3).