Abstract
This study aimed to investigate the pathogenicity of extraintestinal pathogenic Escherichia coli (ExPEC) isolated from dog and cat lung samples in South Korea. A total of 101 E. coli isolates were analyzed for virulence factors, phylogroups, and O-serogroups, and their correlation with bacterial pneumonia-induced mortality was elucidated. P fimbriae structural subunit (papA), hemolysin D (hlyD), and cytotoxic necrotizing factor 1 (cnf1) were highly prevalent in both species, indicating correlation with bacterial pneumonia. Phylogroups B1 and B2 were the most prevalent phylogroups (36.6% and 32.7%, respectively) and associated with high bacterial pneumonia-induced mortality rates. Isolates from both species belonging to phylogroup B2 showed high frequency of papA, hlyD, and cnf1. O-serogrouping revealed 21 and 15 serogroups in dogs and cats, respectively. In dogs, O88 was the most prevalent serogroup (n = 8), and the frequency of virulence factors was high for O4 and O6. In cats, O4 was the most prevalent serogroup (n = 6), and the frequency of virulence factors was high for O4 and O6. O4 and O6 serogroups were mainly grouped under phylogroup B2 and associated with high bacterial pneumonia-induced mortality. This study characterized the pathogenicity of ExPEC and described the probability of ExPEC pneumonia-induced mortality.
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Introduction
Escherichia coli is the most common gram-negative bacterium in the gastrointestinal tract and causes extraintestinal infections in humans and animals, including urinary tract infections (UTIs), neonatal meningitis, prostatitis, and pneumonia1,2,3. E. coli strains that cause diseases other than those of the digestive organs are referred to as extraintestinal pathogenic E. coli (ExPEC)4,5. The pathogenicity of ExPEC is phylogenetically and epidemiologically different from that of commensal and intestinal E. coli strains. Previous reports showed that ExPEC strains contain specific virulence factor encoding genes: cytotoxic necrotizing factor 1 (cnf1), hemolysin D (hlyD), P fimbriae structural subunit (papA), S and F1C fimbriae subunits (sfa/foc), A firmbrial adhesins (afa), group 2 capsular polysaccharide units (kps MII), and aerobactin receptor (iutA)1,6.
Previous studies established the pathogenesis of ExPEC-induced UTIs7,8, meningitis9, sepsis10,11, and pyometra12 in various species. However, limited cases and analyses of ExPEC-induced pneumonia have been reported worldwide. E. coli isolates from the lungs of cats contained various virulence factor encoding genes, including cnf1, papA, hlyD, and kps MII6,13. In addition, E. coli isolates from dogs with pneumonia expressed cnf1, papA, kps MII, and sfaS14,15. We previously reported that E. coli isolated from the lungs of a dog with acute pneumonia in South Korea contained CNF1s and hemolysins. This was categorized as a case of ExPEC pneumonia-induced mortality in the absence of other infections16. However, there are limited number of ExPEC cases to establish the correlation between pneumonia and ExPEC, particularly the correlation between the frequency of occurrence of virulence factors and bacterial pneumonia-induced mortality.
This study aimed to investigate the pathogenicity and characteristics of ExPEC isolated from dog and cat lung samples in South Korea. The results established the correlation between bacterial pneumonia and various characteristics of ExPEC.
Results
Detection of virulence factors in E. coli isolates
PCR analysis of the 101 E. coli isolates from dog and cat lung samples revealed the presence of virulence factors (Table 1). The virulence factor hlyD was prevalent in the E. coli isolates from both dogs and cats (30.3% and 77.1%, respectively). The overall frequency of occurrence of cnf1 in both species was 36.6%, with a relatively lower species-specific frequency of occurrence of 18.2% in dogs and 71.4% in cats. The virulence factor iutA was detected at a low frequency of 14.3% in cats and 34.8% in dogs. Other factors like papA, kps MII, and focG exhibited an overall frequency of occurrence of 33.7%, 30.7%, and 21.8%, respectively. The overall frequency of occurrence of sfa was 8.9% (species-specific frequency, 4.5% in dogs and 17.1% in cats). Virulence factors stx1, stx2, eae, and afa were not detected in any isolate (Table 2). Histopathological examination revealed that bacterial pneumonia-induced mortality was 28.7% (species-specific mortality rate, 25.8% in dogs and 34.3% in cats) (Table 2). The frequency of occurrence was high in papA (15/29 cases), hlyD (18/29 cases), and cnf1 (15/29 cases), and low in iutA (4/29 cases) according to bacterial pneumonia-induced mortality, although there was no significant differences (P > 0.05). The frequency of occurrence of virulence factors focG, kps MII, papA, hlyD and cnf1 was significantly higher in cats than in dogs (P < 0.05). E. coli isolates from lung samples of cats showed no significant differences in virulence factor distribution between cases of bacterial pneumonia-induced mortality and mortality due to other causes. However, in dogs, the distribution of papA (7/17 cases, 41.18%), hlyD (10/17 cases, 58.82%) and cnf1 (7/17 cases, 41.18%) was significantly higher in E. coli isolates from cases of bacterial pneumonia-induced mortality than mortality due to other causes (P < 0.05).
Correlation between the phylogroups and virulence factors of E. coli isolates
Phylogenetic analysis revealed that 36.6% of E. coli isolates belonged to the phylogroup B2, 32.7% to group B1, 9.9% to group C, 8.9% to group A, 4.0% to group C, 3.0% to group F, 1.0% to group E, and unclassified isolates accounted for 4.0% of the total isolates (Table 3). Most isolates from cases of bacterial pneumonia-induced mortality were classified under phylogroups B1 and B2, while three cases were grouped under phylogroup C, D, and unclassified (one case in each group). In dogs, all phylogroups, including the unclassified group, were identified, and group B1 was the most prevalent (42.4%). Nine deaths due to bacterial pneumonia were reported in the B1 group, and focG, hlyD, and iutA were detected in these cases. The prevalence of phylogroup B2 in dogs was lower than the prevalence of group B1, but mortality due to bacterial pneumonia was detected more in phylogroup B2 (7/12 cases, P < 0.05), and all virulence factors apart from iutA were identified in this group. All isolates from dog lung samples, belonging to phylogroup B2 contained the virulence factor-encoding genes papA, hlyD, and cnf1. In isolates from dog lung samples belonging to phylogroups A, C, D, E, and F, mortality due to bacterial pneumonia was not established, and one case was grouped under the unclassified phylogroup. In isolates from cat lung samples, phylogroups B2 (71.4%), B1 (14.3%), C (11.4%), and D (2.9%) were identified. The most prevalent phylogroup was B2, and hlyD and cnf1 were detected in most E. coli isolates from cat lung samples. Moreover, it was observed that in the E. coli isolates belonging to this B2 group, other virulence factors, including papA, kps MII, and focG were detected. In one of the two cases (from cat lung samples) of death due to bacterial pneumonia belonging to phylogroup B1, virulence factors papA, sfaS, hlyD, and iutA were detected, while the presence of iutA was confirmed in the other case. However, virulence factors were not identified in the mortality cases grouped under phylogroup C, and only focG was detected in one bacterial pneumonia-induced mortality case grouped under phylogroup D.
Correlation between the O-serogroups and virulence factors of E. coli isolates
O-serogrouping of all isolates was performed and the frequency of occurrence of different virulence factors in the identified serogroups was evaluated (Table 4). In this study, 28 O-serogroups were identified, and 13 isolates contained unidentified serogroups (Table 4). Nine O-serogroups (O4, O6, O7, O8, O25, O29, O54, O88, and O128) were detected in both dogs and cats, with 13 groups identified from dog lung samples (O9, O11, O36, O41, O60, O78, O81, O89, O91, O131, O156, O161, and O166) and six from cat lung samples (O2, O22, O51, O56, O83, and O102). The most prevalent O-serogroup in dogs was O88 (n = 8), followed by O6 (n = 7), O8 (n = 6), O4 (n = 5), O89 (n = 4), O128 (n = 3), and O9 (n = 3). The detection of virulence factors was the highest in the O6 and O4 serogroups identified from dog lung sample isolates. Six isolates belonging to serogroup O6 were found to express kps MII, papA, hlyD, and cnf1, and all isolates grouped under serogroup O4 expressed papA, hlyD, and cnf1. Bacterial pneumonia-induced mortality was most prevalent in serogroup O4 (4 cases, P < 0.05), followed by O36 (3 cases), O54 (2 cases), and O6 (2 cases). In cat lung sample isolates, serogroup O4 was the most prevalent (6 cases), followed by O6 (5 cases), O8 (4 cases), O51 (4 cases) and O25 (3 cases), although there was no statistical association. All isolates belonging to serogroup O4 expressed papA, hlyD, and cnf1, and two cases of bacterial pneumonia-related death were reported. In all isolates belonging to serogroups O6, O25, and O51, apart from one case, kps MII, papA, hlyD, and cnf1 were detected. In isolates grouped under serogroup O25, bacterial pneumonia-induced mortality was not reported. However, bacterial pneumonia-induced mortality was reported in isolates belonging to serogroups O6 (2 cases) and O51 (1 case). Only hlyD was detected in isolates from serogroup O8, and one case of bacterial pneumonia-induced death was confirmed in this serogroup. The correlation between phylogenetic groups and O-serogroups is shown in Table 5. Apart from phylogroup C of serogroup O8 (n = 4, dog lung samples and n = 3, cat lung samples) and phylogroup A of serogroup O89 (n = 3, dog lung samples), most E. coli isolates from dog and cat lung samples were classified under phylogroups B1 and B2, irrespective of the assigned O-serogroups.
Discussion
Cases of ExPEC infections, including UTIs and meningitis, have been reported in various species. However, there is limited information on ExPEC-induced pneumonia. We previously reported suspected cases of ExPEC-induced pneumonia in dogs16. In addition, cases of E. coli infection in dog20 and cat6 lungs have also been reported, and the virulence factors associated with ExPEC are different from those linked to commensal or intestinal pathogenic E. coli strains4. Moreover, since E. coli is a primary infectious agent that decreases immunity to secondary infections by other pathogens21, the investigation of E. coli respiratory infections is imperative. To gain more insight into the characteristics of ExPEC-induced pneumonia in dogs and cats, the prevalence and pathogenicity of E. coli isolates from dog and cat lung samples were analyzed.
The present study described the characteristics of E. coli isolated from the lungs of dogs and cats that died due to bacterial pneumonia. The virulence factors papA, hlyD, and cnf1 were relatively prevalent and exhibited differences in the frequency of their occurrence in pneumonia-induced mortality cases. A major virulence factor in ExPEC is α-hemolysin, which causes cell lysis and damage12,22. The hlyCABD operon codes for α-hemolysin, and hlyD participates in the secretion of α-hemolysin23. ExPEC infections are characterized by the presence of virulence factors like cnf, which causes apoptosis, and papA24,25. The hly virulence factor is present in the chromosomes of pathogenicity islands and is associated with other virulence factors, including pap and cnf23,26, indicating that these virulence factors are expressed together. Previous studies showed that ExPEC isolates from pneumonia-infected cats6,13 and dogs15 express hly, cnf1, and papA. In addition, lung injury was found to be severe in hly- and cnf-positive ExPEC strains in a rat pneumonia model27. The high frequency of occurrence of papA, hlyD, and cnf1 in the present study suggests that E. coli strains, which possess these virulence factors, can cause bacterial pneumonia in dogs and cats, and may potentially lead to pneumonia-associated mortality.
The presence of kps MII, a virulence factor-encoding gene that also encodes for the K2 capsule protein, is a characteristic feature of neonatal meningitis caused by E. coli. It protects E. coli from phagocytosis and complement-mediated death28,29. In the present study, the E. coli isolates from neither species showed a significant association with the development of bacterial pneumonia, which may be attributed to E. coli viability. In the present study, afa was not detected in any E. coli isolates. This gene has been identified in uropathogenic E. coli (UPEC) and ExPEC strains but not in commensal strains of E. coli, which may indicate that it is not gastrointestinal-derived E. coli30,31. An earlier study showed that only the UTI isolates expressed afa from a total of 40 E. coli isolate samples collected from dogs and cats32, indicating that afa may not be highly prevalent in ExPEC strains. The adherence factors Foc and SfaS help the pathogen to bind to specific host receptors33,34. An earlier study showed that sfa-foc adhesion occurs in specific cell types, indicating differences in colonization depending on the host35. Therefore, in the present study, it is expected that the attachment of E. coli in the lung samples collected from dogs and cats was correlated with other adhesion molecules. The factor iutA engages in iron uptake, which is essential for colonization and bacterial growth in the host and becomes virulent in certain strains36,37. According to Landgraf et al.37, iutA is not essential for UPEC strains. In the present study, iutA was identified in some E. coli isolates from both species, and the frequency of occurrence of this gene was higher in the dog sample isolates than in the lung samples collected from cats. However, iutA did not correlate with bacterial pneumonia-induced mortality.
Shiga toxin 1 (stx1) and stx2 are the primary virulence factors of Shiga toxin-producing E. coli (STEC) strains, which cause diarrhea post-intestinal infection. In addition, intimin, a bacterial adhesion molecule that regulates intestinal epithelial cell adhesion of STEC strains, is encoded by eae38. The results from this study showed that stx1, stx2, and eae were not detected in the virulence factor analysis of E. coli isolates collected from the lung samples of dogs and cats. A previous study showed differences in E. coli isolates from fecal and lung samples14. This suggests that E. coli isolates from the lungs were derived from extraintestinal sources, and the possibility of contamination by intestinal E. coli after death or during autopsy20 can therefore be excluded.
In this study, bacterial pneumonia-induced mortality was higher in cats (34.3%) than in dogs (25.8%). Although more than 60% of E. coli isolates from cat lung samples contained papA, hlyD, and cnf1, no significant association between these virulence factors and mortality due to bacterial pneumonia, similar to the correlation established in the isolates from dog lung samples, could be established in cats. Most dogs used in this study were raised by humans as companion animals (55/66 cases), while more than half of the cats (19/35 cases) included in this study (data not shown) were strays. There is a possibility of differences in E. coli pathogenicity based on differences in the environment that these two species were exposed to. Earlier studies showed that E. coli strains infecting different species were similar, especially strains infecting humans and companion animals, and the possibility of inter-species E. coli transmission was suggested39,40,41. On the other hand, wild animals were infected with E. coli strains different from the strains infecting humans and other animals, and the infection was dependent on the species and its habitat35,42. Thus, the present study showed that the frequencies of occurrence of virulence factors in E. coli isolates from dogs and cats were different and were linked to differences in their habitats.
In this study, the phylogroups B1 (32.7%) and B2 (36.6%) were detected relatively more than the other phylogroups, and the mortality due to bacterial pneumonia was higher in these phylogroups (11/29 and 15/29 cases, respectively). The results showed that the virulence factors papA, hlyD, and cnf1, which are associated with bacterial pneumonia, were highly prevalent in phylogroup B2. Among the E. coli isolates from dogs and cats that died of bacterial pneumonia belonging to phylogroup B2, all seven isolates from dogs (7/7 cases) and seven out of eight isolates from cats (7/8 cases) were confirmed to contain these three virulence factors. E. coli isolates were mainly categorized into four phylogroups: A, B1, B2, and D. The commensal strains of E. coli mostly belong to phylogroups A and B11,4, whereas the ExPEC strains belong to phylogroups B2 and D3,14,15. Previous studies showed that E. coli strains belonging to phylogroups B2 and D caused pathogenic symptoms due to the presence of various virulence factors43. In addition, most ExPEC strains, especially the strains causing UTIs, belong to phylogroups B2 and D and were characterized by the presence of virulence factors that may lead to extraintestinal infections1,14,15,32. Diseases caused by ExPEC strains expressing virulence factors, including papA, hlyD, and cnf1, in cats44, dogs8, and humans45 have been reported. The results from this study were consistent with those from earlier studies and established the correlation between E. coli strains belonging to phylogroup B2 that expressed virulence factors (papA, hlyD, and cnf1) and bacterial pneumonia-induced mortality.
Among the E. coli isolates from dog lung samples belonging to phylogroup B1, nine deaths were caused by bacterial pneumonia, but no correlation with the frequency of virulence factors could be established. Bacterial coinfections and secondary infections caused by E. coli are commonly associated with multiple pathogens, as well as primary infections21,46. Viral and bacterial coinfections have been commonly identified in humans47. An earlier study showed that the mortality rate increased as a result of severe lung lesions caused by coinfection by influenza and E. coli in a mouse model when compared to infections caused by a single pathogen48. The nature of E. coli isolates in the present study (primary or secondary infections) was not established. Nevertheless, it can be hypothesized that E. coli strains belonging to phylogroup B1 with a low frequency of occurrence of virulence factors may enhance pneumonia symptoms due to coinfection with other pathogens.
In this study, 22 and 15 O-serogroups were identified in the isolates from dog and cat lung samples, respectively. The O-antigen is an important virulence factor that is regulated by the repeating polysaccharide chain present in the outer membrane composed of lipopolysaccharides46,49. O-antigen subunits showed variability across different strains containing 181 O-antigens and promoted immune suppression associated with serum sensitivity, which is essential for pathogen survival and outbreak and epidemiological investigations49,50,51. In the present study, the O-serogroup patterns were different in the E. coli isolates from dog and cat lung samples; however, O4 and O6 serogroups were common in both species. Moreover, most isolates belonging to the serogroups O4 and O6 contained various virulence factors, including papA, hlyD, and cnf1. The results of the comparison between O-serogroup and virulence factors of two species showed E. coli isolates from lung samples of dogs belonging to serogroup O4 had significantly higher at bacterial pneumonia-induced mortality than other serogroups. In previous studies, various O-serogroups have been identified in ExPEC infections, and some O-serogroups were more prevalent in certain species. The serogroups O25, O2, O6, and O1 in patient samples were characterized by the presence of various virulence factors51,52,53. Isolated ExPEC samples from dogs and cats were grouped into serogroups O4 and O6 and characterized by the presence of various virulence factors1,54, specifically samples isolated from pneumonia-infected lungs of dogs15,20 and cats6 containing papA, hlyD, and cnf1. The results of comparison between O-serogroups and virulence factors from this study showed that serogroup O4 and O6 were relatively prevalent than other serogroups for virulence factors papA, hlyD, and cnf1 with bacterial pneumonia in dogs and cats, although there was no significant association with bacterial pneumonia-induced mortality, except for serogroup O4 in dogs. Other O-serogroups have also been reported in addition to the dominant serogroups. According to Yuri et al.54, serogroups O25, O11, and O75 are commonly prevalent serogroups detected in canine UTIs. In human ExPEC cases, O18 associated with sfaS and cnf1 expression in meningitis1, and O11, O17, and O77 associated with papA, iutA, and kpsM II expression55 have been reported. In the present study, in addition to the dominant O4 and O6 serogroups, other O-serogroups were also identified, which may be associated with ExPEC pathogenicity. The relationship analysis between phylogroups and O-serogroups showed that most isolates belonged to serogroups O4 and were categorized under phylogroup B2. According to previous studies, pathogenic O-serogroups are associated with phylogroup B2 that is characterized by the presence of various virulence factors56,57. Serogroups O4 and O6 were not correlated with bacterial pneumonia since E. coli isolates were not concentrated in specific O-serogroups and various O-serogroups were identified. Most isolates belonging to serogroup O4 and O6 were classified under phylogroup B2 that is associated with bacterial pneumonia, suggesting a possible correlation between bacterial pneumonia and frequency of occurrence of virulence factors.
In conclusion, this study showed that ExPEC can cause pneumonia in dogs and cats and is an important pathogen implicated in the death of animals depending on pathogenicity. Pathogenicity analysis confirmed the correlation between frequency of occurrence of virulence factors and bacterial pneumonia-induced mortality, and this association was observed in specific ExPEC strains that contained the virulence factor-encoding genes papA, hlyD, and cnf1. In addition, bacterial pneumonia-induced mortality in dogs is more likely to be associated with strains belonging to phylogroups B2 and O-serogroups O4 and O6, and although there were no significant differences in association, strains belonging to phylogroups B2 and O-serogroups O4 and O6 were also dominant in cats which died from bacterial pneumonia. The results of this study described the prevalence and pathogenicity of ExPEC in lung samples collected from dogs and cats in South Korea and provide insights into the correlation between ExPEC strains and bacterial pneumonia.
Materials and methods
E. coli isolates
From January 2020 to June 2022, 101 lung samples from 66 dogs and 35 cats (companion and stray dogs and cats) that died in animal hospitals and outdoors were collected. The sample collection date and location were recorded using application documents. All lung samples were submitted to the Animal and Plant Quarantine Agency (APQA, South Korea) for diagnosis of gross and histopathological lesions. Following the APQA standard autopsy guidelines, samples with bacterial lesions of the lungs, including fibrinous and suppurative pneumonia, pleurisy, and bacterial colonies were identified but no lesions were identified in other organs and were declared death due to bacterial pneumonia. After the autopsy, all samples were stored at 4 ℃ until further experimentation. Samples were first streaked onto MacConkey (MAC; BD, Franklin Lakes, NJ, USA) and sheep blood agar (BA; Asan, Korea), and then incubated in an atmosphere of 5% CO2 at 37 ℃ for 15–18 h. Pure colonies were transferred to a new BA plate and incubated under similar conditions for the same duration. Pure colonies were then isolated, and bacterial species were identified using the VITEK 2 system (bioMérieux, Craponne, France). All E. coli isolates were stored using the BRIX Microvials system (BASIC SCIENCE, Korea) at − 80 ℃ until further experimentation.
Determination of occurrence frequency of virulence factors and phylogenetic analysis
Genomic DNA was extracted using the boiling method. Specific primers for the virulence factors used in this study are listed in Table 1. Amplification was performed in a thermocycler (Takara, Shiga, Japan) with specific cycling conditions. The cycling conditions for focG, kps MII, papA, sfaS, afa, hlyD, and iutA were as follows: an initial cycle at 95 ℃ for 15 min, 35 amplification cycles consisting of denaturation at 95 ℃ for 30 s, annealing at 60 ℃ for 30 s, followed by extension at 72 ℃ for 30 s, and a final cycle of extension at 72 ℃ for 10 min17. The cycling conditions for stx1, stx2, eae, and cnf1 are as follows: an initial cycle at 94 ℃ for 5 min, 35 to 40 amplification cycles consisting of denaturation at 94 ℃ for 1 min, annealing at 55 ℃ for 1 min, followed by extension at 72 ℃ for 1 min, and a final cycle of extension at 72 ℃ for 10 min18,19. Phylogenetic analysis of the 101 E. coli isolates was performed based on PCR analysis of chuA, yjaA, arpA, TspE4.C2, and trpA, according to the previously established procedure58. Based on the PCR results, each isolate was assigned to a specific phylogenetic group (A, B1, B2, C, D, E, and F).
O-serogrouping
O-serogroups were determined using PCR, according to the previously described procedure59. E. coli isolates were cultured on BA at 37 ℃ overnight. Genomic DNA was extracted by boiling the cultured colonies. O-genotyping multiplex PCR was performed using primer sets MP-1–MP-20.
Statistical analysis
All results are expressed as percentage of isolates and presence of virulence factors. The frequency of occurrence of virulence factors in E. coli isolates from dog and cat lung samples was statistically compared using the chi-squared test or Fisher’s exact test followed by Holm’s post-hoc test. The P-value was calculated, and statistical significance was set at P < 0.05.
Ethics approval and consent to participate
Informed consent was obtained from all dog and cat owners involved in the study.
Data availability
The data presented in this study are available on request from the corresponding author.
References
Smith, J. L., Fratamico, P. M. & Gunther, N. W. Extraintestinal pathogenic Escherichia coli. Foodborne Pathog. Dis. 4(2), 134–163. https://doi.org/10.1089/fpd.2007.0087 (2007).
Poolman, J. T. & Wacker, M. Extraintestinal pathogenic Escherichia coli, a common human pathogen: Challenges for vaccine development and progress in the field. J. Infect. Dis. 213(1), 6–13. https://doi.org/10.1093/infdis/jiv429 (2016).
Flament-Simon, S. C. et al. Molecular characteristics of extraintestinal pathogenic E. coli (ExPEC), uropathogenic E. coli (UPEC), and multidrug resistant E. coli isolated from healthy dogs in Spain. Whole genome sequencing of canine ST372 isolates and comparison with human isolates causing extraintestinal infections. Microorganisms 8(11), 1712. https://doi.org/10.3390/microorganisms8111712 (2020).
Russo, T. A. & Johnson, J. R. Proposal for a new inclusive designation for extraintestinal pathogenic isolates of Escherichia coli: ExPEC. J. Infect. Dis. 181(5), 1753–1754. https://doi.org/10.1086/315418 (2000).
Beutin, L. Escherichia coli as a pathogen in dogs and cats. Vet. Res. 30(2–3), 285–298 (1999).
Highland, M. A. et al. Extraintestinal pathogenic Escherichia coli-induced pneumonia in three kittens and fecal prevalence in a clinically healthy cohort population. J. Vet. Diagn. Investig. 21(5), 609–615. https://doi.org/10.1177/104063870902100504 (2009).
Manges, A. Escherichia coli and urinary tract infections: The role of poultry-meat. Clin. Microbiol. Infect. 22(2), 122–129. https://doi.org/10.1016/j.cmi.2015.11.010 (2016).
Valat, C. et al. Pathogenic Escherichia coli in dogs reveals the predominance of ST372 and the human-associated ST73 extra-intestinal lineages. Front. Microbiol. 11, 580. https://doi.org/10.3389/fmicb.2020.00580 (2020).
Kim, K. S. Human meningitis-associated Escherichia coli. EcoSal Plus 7(1), 1. https://doi.org/10.1128/ecosalplus.ESP-0015-2015 (2016).
Mora-Rillo, M. et al. Impact of virulence genes on sepsis severity and survival in Escherichia coli bacteremia. Virulence 6(1), 93–100. https://doi.org/10.4161/21505594.2014.991234 (2015).
Daga, A. P. et al. Escherichia coli bloodstream infections in patients at a university hospital: Virulence factors and clinical characteristics. Front. Cell. Infect. Microbiol. 9, 191. https://doi.org/10.3389/fcimb.2019.00191 (2019).
Melo, R. T. et al. Phylogeny and virulence factors of Escherichia coli isolated from dogs with pyometra. Vet. Sci. 9(4), 158. https://doi.org/10.3390/vetsci9040158 (2022).
Brooks, J. W., Roberts, E. L., Kocher, K., Kariyawasam, S. & DebRoy, C. Fatal pneumonia caused by extraintestinal pathogenic Escherichia coli (ExPEC) in a juvenile cat recovered from an animal hoarding incident. Vet. Microbiol. 167(3–4), 704–707. https://doi.org/10.1016/j.vetmic.2013.08.015 (2013).
Turchetto, S. et al. Phenotypic features and phylogenetic background of extraintestinal hemolytic Escherichia coli responsible of mortality in puppies. Vet. Microbiol. 179(1–2), 126–130. https://doi.org/10.1016/j.vetmic.2015.03.004 (2015).
Moore, M. E. et al. Molecular characteristics, ecology, and zoonotic potential of Escherichia coli strains that cause hemorrhagic pneumonia in animals. Appl. Environ. Microbiol. 87(23), e01471-e1521. https://doi.org/10.1128/AEM.01471-21 (2021).
Kim, G. et al. Fatal pneumonia caused by extraintestinal pathogenic Escherichia coli in a young dog. Korean J. Vet. Re. 62(1), 1–5. https://doi.org/10.14405/kjvr.20210043 (2022).
Franz, E., Veenman, C., van Hoek, A. H., Husman, A. R. & Blaak, H. Pathogenic Escherichia coli producing extended-spectrum β-lactamases isolated from surface water and wastewater. Sci. Rep. 5(1), 1–9. https://doi.org/10.1038/srep14372 (2015).
Clermont, O. et al. Animal and human pathogenic Escherichia coli strains share common genetic backgrounds. Infect. Genet. Evol. 11(3), 654–662. https://doi.org/10.1016/j.meegid.2011.02.005 (2011).
Tóth, I., Hérault, F., Beutin, L. & Oswald, E. Production of cytolethal distending toxins by pathogenic Escherichia coli strains isolated from human and animal sources: Establishment of the existence of a new cdt variant (type IV). J. Clin. Microbiol. 41(9), 4285–4291. https://doi.org/10.1128/JCM.41.9.4285-4291.2003 (2003).
Handt, L. K. et al. Clinical and microbiologic characterization of hemorrhagic pneumonia due to extraintestinal pathogenic Escherichia coli in four young dogs. Comp. Med. 53(6), 663–670 (2003).
Manohar, P. et al. Secondary bacterial infections during pulmonary viral disease: Phage therapeutics as alternatives to antibiotics? Front. Microbiol. 11, 1434. https://doi.org/10.3389/fmicb.2020.01434 (2020).
Bakás, L., Veiga, M. P., Soloaga, A., Ostolaza, H. & Goñi, F. M. Calcium-dependent conformation of E. coli α-haemolysin. Implications for the mechanism of membrane insertion and lysis. Biochim. Biophys. Acta Biomembr. 1368(2), 225–234. https://doi.org/10.1016/S0005-2736(97)00181-8 (1998).
Velasco, E. et al. A new role for Zinc limitation in bacterial pathogenicity: Modulation of α-hemolysin from uropathogenic Escherichia coli. Sci. Rep. 8(1), 1–11 (2018).
Uhlin, B. E., Norgren, M., Båga, M. & Normark, S. Adhesion to human cells by Escherichia coli lacking the major subunit of a digalactoside-specific pilus-adhesin. Proc. Natl. Acad. Sci. 82(6), 1800–1804. https://doi.org/10.1073/pnas.82.6.1800 (1985).
Thomas, W. et al. Cytotoxic necrotizing factor from Escherichia coli induces RhoA-dependent expression of the cyclooxygenase-2 gene. Infect. Immun. 69(11), 6839–6845. https://doi.org/10.1128/IAI.69.11.6839-6845.2001 (2001).
Blum, G., Falbo, V., Caprioli, A. & Hacker, J. Gene clusters encoding the cytotoxic necrotizing factor type 1, Prs-fimbriae and α-hemolysin form the pathogenicity island II of the uropathogenic Escherichia coli strain J96. FEMS Microbiol. Lett. 126(2), 189–195. https://doi.org/10.1111/j.1574-6968.1995.tb07415.x (1995).
Russo, T. A. et al. E. coli virulence factor hemolysin induces neutrophil apoptosis and necrosis/lysis in vitro and necrosis/lysis and lung injury in a rat pneumonia model. Am. J. Physiol. Lung Cell. Mol. Physiol. 289(2), 207–216. https://doi.org/10.1152/ajplung.00482.2004 (2005).
Johnson, J. R. & O’Bryan, T. T. Detection of the Escherichia coli group 2 polysaccharide capsule synthesis gene kpsM by a rapid and specific PCR-based assay. J. Clin. Microbiol. 42(4), 1773–1776. https://doi.org/10.1128/JCM.42.4.1773-1776.2004 (2004).
Wijetunge, D. et al. Characterizing the pathotype of neonatal meningitis causing Escherichia coli (NMEC). BMC Microbiol. 15(1), 1–15. https://doi.org/10.1186/s12866-015-0547-9 (2015).
Keller, R. et al. Afa, a diffuse adherence fibrillar adhesin associated with enteropathogenic Escherichia coli. Infect. Immun. 70(5), 2681–2689. https://doi.org/10.1128/IAI.70.5.2681-2689.2002 (2002).
Qin, X. et al. Comparison of adhesin genes and antimicrobial susceptibilities between uropathogenic and intestinal commensal Escherichia coli strains. PLoS ONE 8(4), e61169. https://doi.org/10.1371/journal.pone.0061169 (2013).
Tramuta, C., Nucera, D., Robino, P., Salvarani, S. & Nebbia, P. Virulence factors and genetic variability of uropathogenic Escherichia coli isolated from dogs and cats in Italy. J. Vet. Sci. 12(1), 49–55. https://doi.org/10.4142/jvs.2011.12.1.49 (2011).
Morschhäuser, J., Hoschützky, H., Jann, K. & Hacker, J. Functional analysis of the sialic acid-binding adhesin SfaS of pathogenic Escherichia coli by site-specific mutagenesis. Infect. Immun. 58(7), 2133–2138. https://doi.org/10.1128/iai.58.7.2133-2138.1990 (1990).
Khan, A. S. et al. Receptor structure for F1C fimbriae of uropathogenic Escherichia coli. Infect. Immun. 68(6), 3541–3547. https://doi.org/10.1128/IAI.68.6.3541-3547.2000 (2000).
Frömmel, U. et al. Adhesion of human and animal Escherichia coli strains in association with their virulence-associated genes and phylogenetic origins. Appl. Environ. Microbiol. 79(19), 5814–5829. https://doi.org/10.1128/AEM.01384-13 (2013).
Chouikha, I., Bree, A., Moulin-Schouleur, M., Gilot, P. & Germon, P. Differential expression of iutA and ibeA in the early stages of infection by extra-intestinal pathogenic E. coli. Microbes Infect. 10(4), 432–438. https://doi.org/10.1016/j.micinf.2008.01.002 (2008).
Landgraf, T. N. et al. The ferric aerobactin receptor IutA, a protein isolated on agarose column, is not essential for uropathogenic Escherichia coli infection. Rev. Lat. Am. Enfermagem 20, 340–345. https://doi.org/10.1590/S0104-11692012000200017 (2012).
Yang, X. et al. Genetic diversity of the intimin gene (eae) in non-O157 Shiga toxin-producing Escherichia coli strains in China. Sci. Rep. 10(1), 1–9. https://doi.org/10.1038/s41598-020-60225-w (2020).
Johnson, J. R. et al. Molecular comparison of extraintestinal Escherichia coli isolates of the same electrophoretic lineages from humans and domestic animals. J. Infect. Dis. 183(1), 154–159. https://doi.org/10.1086/317662 (2001).
Bélanger, L. et al. Escherichia coli from animal reservoirs as a potential source of human extraintestinal pathogenic E. coli. FEMS Immunol. Med. Microbiol. 62(1), 1–10. https://doi.org/10.1111/j.1574-695X.2011.00797.x (2011).
Kidsley, A. K. et al. Companion animals are spillover hosts of the multidrug-resistant human extraintestinal Escherichia coli pandemic clones ST131 and ST1193. Front. Microbiol. 11, 1968. https://doi.org/10.3389/fmicb.2020.01968 (2020).
Murphy, R. et al. Genomic epidemiology and evolution of Escherichia coli in wild animals in Mexico. Msphere 6(1), e00738–e00820. https://doi.org/10.1128/mSphere.00738-20 (2021).
Jeong, Y. W., Kim, T. E., Kim, J. H. & Kwon, H. J. Pathotyping avian pathogenic Escherichia coli strains in Korea. J. Vet. Sci. 13(2), 145–152. https://doi.org/10.4142/jvs.2012.13.2.145 (2012).
Liu, X., Thungrat, K. & Boothe, D. M. Multilocus sequence typing and virulence profiles in uropathogenic Escherichia coli isolated from cats in the United States. PLoS ONE 10(11), e0143335. https://doi.org/10.1371/journal.pone.0143335 (2015).
Kim, D. H. et al. Virulence properties of uropathogenic Escherichia coli isolated from children with urinary tract infection in Korea. Genes Genom. 40(6), 625–634. https://doi.org/10.1007/s13258-018-0664-6 (2018).
Mueller, M. & Tainter, C. R. Escherichia coli. https://www.ncbi.nlm.nih.gov/books/NBK564298/ (2022).
Crotty, M. P. et al. Epidemiology, co-infections, and outcomes of viral pneumonia in adults: An observational cohort study. Medicine 94, 50. https://doi.org/10.1097/MD.0000000000002332 (2015).
Wang, S. et al. Co-infection of H9N2 influenza A virus and Escherichia coli in a BALB/c mouse model aggravates lung injury by synergistic effects. Front. Microbiol. 12, 670688. https://doi.org/10.3389/fmicb.2021.670688 (2021).
Liu, B. et al. Structure and genetics of Escherichia coli O antigens. FEMS Microbiol. Rev. 44(6), 655–683. https://doi.org/10.1093/femsre/fuz028 (2020).
Valvano, M. A. Export of O-specific lipopolysaccharide. Front. Biosci.-Landmark 8(6), 452–471. https://doi.org/10.2741/1079 (2003).
Weerdenburg, E. et al. Global distribution of O-serotypes and antibiotic resistance in extraintestinal pathogenic Escherichia coli (ExPEC) collected from the blood of bacteremia patients across multiple surveillance studies. Clin. Infect. Dis. https://doi.org/10.1093/cid/ciac421 (2022).
Delannoy, S. et al. Characterization of colistin-resistant Escherichia coli isolated from diseased pigs in France. Front. Microbiol. 8, 2278. https://doi.org/10.3389/fmicb.2017.02278 (2017).
Matsumoto, T. et al. Distribution of extraintestinal pathogenic Escherichia coli O-serotypes and antibiotic resistance in blood isolates collected from patients in a surveillance study in Japan. J. Infect. Chemother. 28(11), 1445–1451. https://doi.org/10.1016/j.jiac.2022.07.001 (2022).
Yuri, K., Nakata, K., Katae, H., Tsukamoto, T. & Hasegawa, A. Serotypes and virulence factors of Escherichia coli strains isolated from dogs and cats. J. Vet. Med. Sci. 61(1), 37–40. https://doi.org/10.1292/jvms.61.37 (1999).
Johnson, J. R. et al. Distribution and characteristics of Escherichia coli clonal group A. Emerg. Infect. Dis. 11(1), 141. https://doi.org/10.3201/eid1101.040418 (2005).
Riley, L. Pandemic lineages of extraintestinal pathogenic Escherichia coli. Clin. Microbiol. Infect. 20(5), 380–390. https://doi.org/10.1111/1469-0691.12646 (2014).
Nojoomi, F. & Ghasemian, A. The relation of phylogroups, serogroups, virulence factors and resistance pattern of Escherichia coli isolated from children with septicemia. New Microbes New Infect. 29, 100517. https://doi.org/10.1016/j.nmni.2019.100517 (2019).
Clermont, O., Christenson, J. K., Denamur, E. & Gordon, D. M. The clermont Escherichia coli phylo-typing method revisited: Improvement of specificity and detection of new phylo-groups. Environ. Microbiol. Rep. 5(1), 58–65. https://doi.org/10.1111/1758-2229.12019 (2013).
Iguchi, A. et al. A complete view of the genetic diversity of the Escherichia coli O-antigen biosynthesis gene cluster. DNA Res. 22(1), 101–107. https://doi.org/10.1093/dnares/dsu043 (2015).
Acknowledgements
The authors thanks the owners of the affected dogs and cats for their active cooperation.
Funding
This research was supported by the program (B-1543069-2023-24) of Animal and Plant Quarantine Agency (APQA) and Ministry of Agriculture, Food and Rural Affairs (MARFA).
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Conceptualization, K.L. and B. M.; software, C.S.Y.; validation, B.M., M.H. and S.L.; formal analysis, K.L.; investigation, C.S.Y. and M.H.; resources, S.L.; writing—original draft preparation, C.S.Y.; writing—review and editing, K.L.; funding acquisition, B.K. All authors have read and agreed to the published version of the manuscript.
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Yun, C.S., Moon, BY., Hwang, MH. et al. Characterization of the pathogenicity of extraintestinal pathogenic Escherichia coli isolates from pneumonia-infected lung samples of dogs and cats in South Korea. Sci Rep 13, 5575 (2023). https://doi.org/10.1038/s41598-023-32287-z
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DOI: https://doi.org/10.1038/s41598-023-32287-z
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