UPEC are major cause of urinary tract infections and may be responsible for nearly 90% of UTI
]. There are four main phylogenetic groups (A, B1, B2, and D) of UPEC and the phylogenicity has been reported to play an important role in virulence of these pathogens
]. Although a number of studies on this subject have been carried in various parts of the world, such data is not available from Pakistan where UTI are very common. This study was designed to characterize local isolates of UPEC with respect to phylogenicity and distribution of most important virulence factors. Although our study is comprised of a relatively small number of samples and therefore faces limitations in statistical analysis, it provides important information about the phylogenetic background of uropathogenic E.coli
from this region.
In the present study, we found that 50% of our isolates belonged to a single phylogenetic group B2 which is in line with some other studies
We found that among all phylogenetic groups, most of the virulence related genes were present in significantly higher proportion in phylogenetic group D isolates except papC
gene, which was more frequent in B2 isolates. This is in accordance with the report of Nowrouzian et al.
] who found that most B2 strains carried genes for P-fimbriae.
Our results also supported some previous reports indicating greater association of traditionally recognized uropathogenic virulence factors (e.g. pap
, and hly
) with groups D and B2 as compared with A and B1
Among adhesins, sfaDE
was highly (27%) prevalent among local UPEC isolates followed by papC
(24%) and afaBC3
(8%) genes. We found that frequency of sfa
was higher as compared to other genes. In some of isolates, multiple adhesin genes were identified (pap
) where as in some other isolates these genes were present independently. Here again, phylogenetic group D was most prominent. Our findings are comparable with those of Le Bouguenec et al.
] who found pap
operon in 79.4% of the pyelonephritis strains, either present alone (51.5%) or in association with either afa
operon (27.9%), whereas in 12.4% and 22.7% of the isolates the adhesive properties were associated with the presence of afa
operons, respectively. It is important to note that we did find these three operons in two isolates. The simultaneous presence of pap
, and sfa
operons has not been reported before.
The intimin adhesin (eaeA
gene) is an attachment and membrane damaging pathogenicity factor. It was found in 19% of the isolates. The presence of this adhesin gene has also been previously reported in UPEC
] though it is well known that it is associated with diarrhoeagenic E. coli
Among toxins, hemolysin was identified in 22 (37%) isolates. These findings are in line with those of Bingen-Bidois et al.
] who found hly
gene in 34% of UPEC isolates. In the present study, all hlyA
gene positive isolates hemolysed human erythrocytes.
Our UPEC isolates showed considerable cytotoxic effects and 20% were positive for cnf1
gene. When 27 (46%) UPEC isolates having cnf1
or both genes were used to invade the Vero cells, highest toxicity (80-100% vero cell death) was observed in hlyA
gene positive isolates (n
06), followed by hlyA+cnf1
04) and cnf1
01) isolates. This is in agreement with the reports suggesting that during the early stages of uropathogenesis, hly
plays a major role in the damage of uroepithelium. It has also been reported that in many uropathogenic E. coli
the loci of hly
are often linked and responsible for the severity of urinary tract infections
High cytotoxicity levels were not always found associated with cnf1
positive isolates in this study because some isolates which were positive for other virulence genes, showed cytotoxic effect as well. On the other hand, some isolates with cnf1
did not express any cytotoxic effect on Vero cells. It may be due to an insertion of a thymidine base at position 3325 in the C-terminal domain of cnf1,
which encodes the catalytic region of the toxin
]. The cumulative presence of papC
is the evidence of pathogenicty island IIJ96
, which is highly prevalent among UTI isolates
]. In the present study the simultaneous presence of papC
was observed in 4 UPEC isolates.
Highly hemolytic isolates belonged to phylogenetic group D, while highly cytotoxic isolates belonged to group B2 and D. This is in partial disagreement with the studies of Zhang et al.
] who found that hly
positive isolates mainly belonged to phylogenetic group B2.
It can be concluded that all local UPEC isolates have a battery of virulence factors which makes them a serious and challenging health problem. The simultaneous presence of pap, afa, and sfa operons has been reported for the first time underlining the dynamic nature of these isolates. Another finding of interest was the prominence of Group D in relevance to the presence of virulence factors. The previous reports from other parts of the globe mostly highlight Group B2.