A Comparative Analysis of PCR Results between the Bacteriology Department and the Nursing Home Isolates
Sequence type 131 of E. coli is one of the deadliest groups of pathogens that colonise the gut of human beings and other mammals. The pathogens are known to produce CTX-M-15 ESBL. This pathogenic development has made them resistant and difficult to control (Clermont et al. 274). The clone is particularly resistant to fluoroquinolone, which is one of the most powerful β-lactams found in the market today. For this reason, the clone is associated with high rates of morbidity and mortality in the society.
The ST131 clone has continued to cause health problems in many settings. The major causes of these health vices are attributed to their ability to form ST131 sub-clones H30-R and H30-RX. In order to effectively control these pathogens, researchers need to isolate the ST131 clones from other types of ESBL producing E. coli. The isolates need to be analysed extensively to identify the major factors behind their prevalence and their major characteristics (Foxman 649).
It is also important to identify the pathogens that belong to the H30-RX sub-clone and those associated with sub-clone H30-R. The identification will help to deal with each one separately and effectively.
As stated in the methodology section, the polymerase chain reaction (PCR) was one of the methods used in the study of isolates obtained from the nursing home. The method was used for a number of reasons. For example, previous studies have proven that the technique is effective in conducting research based on DNA sequencing and cloning. It is also effective in the analysis of DNA-based phylogeny. In this case, PCR has been used for phylogenetic and molecular purposes.
It is used to analyse the evolutionary relationships that occur between groups of organisms. It is noted that PCR involves the amplification of DNA fragments. As a result, millions of copies of the parent DNA can be generated from a small sample (Banerjee et al. 6386). Consequently, the technique makes it possible for scientists to conduct studies on a significant amount of DNA that can be easily worked on.
In this study, the main aim of using PCR was to identify the genes present in the isolates obtained from the various sources. The ability of the technique to check for a variety of genes at the same time explains its adoption in various research endeavours in this field.
The presence of various genes tested in this study included strB, aac(6′)Ib-cr, strA, blaCTX-M-15, blaTEM-1B, mph(A), catB3, catA1, sul2, sul1, tet(B), dfrA17, aac(3)-IId, blaOXA-1, aadA5, blaCTX-M-14, blaTEM-1C, tet(A), blaCTX-M-27, nfaE, iha, sat, iss, PrfB, gad, ipfA, SenB, and ireA among others. The PCR results allowed the researcher to detect the presence of sub-clones H30-RX and H30-R (Perez-Perez and Hanson 2153). Furthermore, this technique helped to note the absence of these genes.
The results obtained through the use of PCR method have been used to determine the sub-clone that each of the isolates belongs to. It is important to note that a total of 78 isolates were tested in this study. 64 out of the 78 isolates were obtained from a single nursing home. Only 19 of the 64 isolates tested negative for the ST131 sub-clone H30-RX PCR. The remaining 45 isolates tested positive for ST131 sub-clone H30-RX. The isolates that tested positive for ST131 sub-clone H30-RX PCR made up 70.31 percent of the total isolates collected from the nursing home. The remaining 29.69 percent composed of isolates that tested as ST131 sub-clone H30-RX PCR negative.
The production of ESBLs by the organism makes them resistant to a wide range of antimicrobial agents (Perez-Perez and Hanson 2153). Understanding the characteristics of the sub-clones and root cause of the resistance enables researchers to device better and effective ways of combating the prevalence of the E. coli. It appears that the urine isolates from the Nursing Home contained high lethal effects as the gene of the E. coli was modified to form a different strain.
In essence, a tally of above 50% of the urine extracts complemented the resistant pathogen. Therefore, it can be argued that the outpatients are susceptible to more than 50% pathogenic effect while in the nursing home. In fact, 20% percent of blood cultures collected from this facility proved on its presence within the human blood stream. Probably, the 30% difference was due to the variations caused by the available small sample. If larger samples were available, the affected population might have dictated highly accurate values close to 50%.
The first test results for the study were obtained from the H30-RX PCR. The isolates that tested negative were subjected further to ST131 H30 PCR and their results recorded for interpretation. As stated earlier, ESBLs appears to be the relevant cause for antimicrobial resistance that is observed among E. coli clones and sub-clones.
To this end, the study incorporated results obtained from the ESBL Combination disc test to help in the detection of the enzymes. It is also important to determine the specific enzyme that is present in order to determine ST131 sub-clones present.
The ResFinder 2.1 technique has a great importance in this section. The results obtained from the procedure helps to determine the prevalence of genes that are known to confer antimicrobial resistance to microbes. The VirulenceFinder procedure was also used in the study. The results obtained from the technique help determine the prevalence of the factors that confer virulent characteristics to the isolates.
PCR for H30RX among the Isolates in the Nursing Home
Sequence type 131 of E. coli may colonise the gut of human being and other mammals. The fact that the pathogens produce CTX-M-15 ESBL has made the control strategies ineffective. The clone is particularly resistant to fluoroquinolone, which is among the most efficient β-lactams in the market today (Yokota et al. 52). For this reason, there have been high morbidity and mortality rates today associated with the clone.
The ST131 clone has continued to cause health problems in the communities as well as health centres even after research showed that it led to the production of the ST131 sub-clones H30-R and H30-RX (Nicolas-Chanoine, Xavier, and Madec 550). In order to control these pathogens effectively, researchers need to establish mechanisms for isolating the ST131 clones from other types of ESBL producing E. coli and study them extensively. It is also important to identify which of these pathogens belong to the H30-RX sub-clone and those which belong to sub-clone H30-R.
As stated in the methodology section, the polymerase chain reaction (PCR) was one of the methods used in the study of isolates obtained from the nursing home. The method was used based on its efficiency in conducting researches based on DNA sequencing and cloning, as well as DNA-based phylogeny. In this case, PCR has been used for phylogenetic and molecular purpose, studying the evolutionary relationships that occur between groups of organisms (Foxman 650).
Since PCR involves the amplification of DNA fragments, millions of copies of the parent DNA can be generated. The technique allows scientists to conduct research on a significant amount of DNA that can be easily worked on.
In the current study, the main aim of using PCR was to identify the genes present in the isolates. It ability to check for a variety of genes at ago has seen the technique adopted across the world. Among the genes that were tested for in this study include strB, aac(6′)Ib-cr, strA, blaCTX-M-15, blaTEM-1B, mph(A), catB3, catA1, sul2, sul1, tet(B), dfrA17, aac(3)-IId, blaOXA-1, aadA5, blaCTX-M-14, blaTEM-1C, tet(A), blaCTX-M-27, nfaE, iha, sat, iss, PrfB, gad, ipfA, SenB, as well as, ireA.
Based on the PCR result obtained, the researcher will be in a position to detect the presence of sub-clones H30-RX and H30-R. Through the use of the technique, their absence can also be noted.
The results of the PCR have been used to determine the sub-clone that each of the isolates belongs to. It is important to note that a total of 78 isolates were tested. Of this number, 60 were obtained from urine, 15 from blood culture, and 3 from rectal swabs. 64 out of the 78 isolates used were obtained from a single nursing home.
Only 19 of the 64 isolates were negative for the ST131 sub-clone H30-RX PCR. The rest, 45 isolates tested positive for ST131 sub-clone H30-RX. The isolates that tested positive for ST131 sub-clone H30-RX PCR made up 70.31 percent of the total isolates collected from the nursing home. The remaining 29.69 percent was composed of isolates that tested to be ST131 sub-clone H30-RX PCR negative.
It is important to comparethe CTX-M groupsin the study. As mentioned in the results section:
- CTX-M Group 1= 82% positive of the isolates
- CTX-M Group 9= 18% positive of the isolates
- TEM: 89% were positive for this gene
- SHV: 2% were positive for the gene
- OXA-1: 62% were positive for the gene
Aminoglycosides
- 1-Streptomycin; 76% of isolates were resistance
- 2-Gentamicin; 69% of isolates were resistance
- 3-Kanamycin; 62% of isolates were resistance
- Sulphonamide 94% of isolates were resistance
- Tetracycline: 89% of the isolates were resistance
Fluoroquinolones
- Ciprofloxacin: 99% were resistant.
- Naladixic acid: 100% were resistant.
Beta-lactam antibiotic
- Cefpodoxime; 100 % were resistant
- Cefotaxime; 100% were resistant
- Ampicillin; 100% were resistant
- Ceftazidime; 48% were resistant
CTX-M
The CTX-M enzyme was first discovered in 1995. The enzyme was considered to be of great importance following a rapid increase in its variants among patients suffering from bacterial diseases (Foxman 650). The enzyme has been noted to hydrolyse cefotaxime therefore causing its resistance.
Although the enzyme is highly deactivated by clavulanic acid, it has been found to be susceptible to tazobactam. Presently, research has shown that there exist over 40 variants of the enzyme. The variants have been classified into five major sequence subtypes of amino acids. Isolates of these variant enzymes have been reported in health facilities across Greek especially among E. coli bacterial species (Foxman 650).
Research has also shown that the CTX-M variants are transferable from one bacterial species to another. When the enzyme producing bacterial isolates are mated in agar containing E. coli, they transfer the resistance to the previously susceptible species (Perez-Perez and Hanson 2154). The development of such clones has made it difficult to control the E. coli since they continue to evolve in terms of their antibiotics resistance.
CTX-M-15
The CTX-M-15- type ESBLs have been viewed to have varying resistance to the cephalosporins and other β-lactam antimicrobial agents, such as penicillin and monobactams. Clavulanic acid and sulbactam inhibitors are considered to be rarely effective against these enzymes. Research has however shown that the bacteria are highly susceptible to the tazobactam inhibitor. Most E. coli species producing the ESBL have been cloned from the Enterobacter cloacae Bacterial species (Foxman 649).
Since the enzyme has exhibited great levels of resistance even in cases where antibiotic- inhibitor combinations have been used, researchers have resulted to the manufacture of antimicrobial agents which has a core that is not made of β-lactam compounds.
Comparison between the ST131 and its H30-RX and H30 Sub-Clones Results andFindings from Other Literature Material
From the literature review, it was indicated that the Escherichia coli sequence type 131 brings about a number of infections. In the current study, the ST131 is perceived as the main cause for a host of extra intestinal infections. A separate study carried out by M’Zali et al. suggest that the ST131 is further responsible for the fluoroquinolone resistance (882). The same clone is also responsible for the production of the extended-spectrum beta-lactamase. However, to understand how each of these medical conditions result the determination of the ST131 and all its sub-clones’ prevalence is necessary.
The results of the current study indicate that, out of all the 88 isolates, the prevalence of the H30-RX stood at 59%. With respect to the H30 sub-clones, the 35 isolates tested had a prevalence of 60%. According to Paterson and Bonomon, the prevalence of such sub-clones has a direct impact on the activity of the drugs that would be used to contain the effects of the E. coli (657).The prevalence of blaTEM-1B , blaCTX-M15, dfrA17, sul 1, and aadA5 antibiotic resistant genes amongst the isolates is indicated in figure 3.5.1.
Similar studies on the same subject have been carried out in to evaluate the gene resistance. In a study by Tamang et al., an extensive molecular report of plasmid-mediated ESBLs lactamases in E. coli strains is reported (17).The samples were obtained from dogs in the Republic of Korea. A total of six hundred and twenty eight E. coli isolates were examined. Out of these, 12 tested positive for CTX-M-type ESBLs. In addition, 22 tested positive for AmpCs.
In the study, a similar incidence of CTX-M-type ESBLs was noted. The group was noted in E. coli samples obtained from dogs and cats. The second sample group was from the United States. According to Tamang et al., the results from the second population group indicated that the CTX-M had a higher frequency of pAmpCs (18).
Results from the Europe station indicated that resistance to ESC among the canine and feline E. coli isolates was equal to the number of genes. However, from the station in China, a high prevalence of blaCTX-M genes was reported. The findings from the current study, as well as the ones reported by Tamang et al., indicate that prevalence varies with location (15). Consequently, addressing the mutation of the E. coli resistance to various compounds require scientist to factor in the location.
Depending on the nature of the compound used to contain the effects of E. coli, the prevalence rates vary. In the study by Tamang et al., the high prevalence was attributed to the high dosage of cephalosporins in the samples obtained from the Republic of Korea (16). Cephalexin is the most common cephalosporin that is used in animal hospitals in the Republic of Korea (Tamang et al. 15). The ESBL-producing canine E. coli isolates contained a total of 6blaCTX-M variants have been reported previously for E. coli strains isolated from companion animals.
The presence of blaCTX-M-14 in an E. coli strain isolated has been discovered in various countries particularly in dogs. According to Tamang et al., the first ever recording of a clinically sick dog was discovered in 2005 in the Republic of Korea (17). The pAmpC gene encoding CMY-2 operates in conjunction with TEM-1. Tamang et al. indicate that the same has been found to be responsible for the E. coli strains isolated from dogs from Europe and the United States (18).
According to Tamang et al., CMY-2 has not been reported before amongdogs inKorea (14). In addition, blaDHA-1 gene was observed in two samples in the study by Tamang et al. (14). Earlier, the prevalence of this gene was recorded among human clinical E. coli isolates. The isolates were from the Republic of Korea. No cases of E. coli strains from companion animals had been recorded in the world before (Tamang et al. 14).
The research was a pioneering investigation into the incidences of blaDHA-1in E. coli samples obtained from canines. The results are very significant. They indicate that there are different forms of ESBL genesin E. coli strains among dogs. The same is an obvious public health concern. Majority of isolates in the current study recorded unique trends with regards toblaCTX-MandblaCMY-2 genes. The two were passed on through conjugation to another E. coli species.
Interestingly, the blaDHA-1 and/or blaTEM-1gene did not transfer to recipient strains. According to M’Zali et al., such a scenario implies that the genetic determinants are different (882). They are to be found in a plasmid that varies from the one bearing the sub-clone genetic material (M’Zali et al. 882).
A study by Mora et al. sought to ascertain whether or not the sub-clone H30-Rx is responsible for the epidemic of ERLP E. coli (6). The results of this study confirm that clonal expansion is the major reason behind the production of CTX-M-15 and fluoroquinolone resistance among E. coli ST131 (Mora et al. 6).
With respect to the current study, the CTX-M had two groups where the positivity to the isolates was examined. In both cases there was a high prevalence of the isolates. As indicated, the CTX-M group 1 recorded 82% of the Isolates while the CTX-M recorded 18% of the isolates. The implication is that, indeed, the sub-clones of the E. coli is responsible for the high resistance from the corresponding drugs.
The resistance exhibited by the sub-clones is brought about by the respective mechanisms of gene transfer. The findings of a study by Lim et al. indicate that a majority of the determinants for both of the said conditions traits are brought about by a horizontal gene transfer (139). With respect to the fluoroquinolone, the resistance was brought about courtesy of a phenomenon described as independent mutation. The current study also had a similar behaviour among the sub-clones.
The difference between the two studies with regards to the resistance levels is such that in the research by Lim et al., the whole-genome SNP-based phylogenies belonged to H30-R (140). In the current study, the resistance was derived from H30- Rx sub-clone.
The H30-R sub-clone is often derived from a single common ancestor. According to DeVincent and Reid-Smith, the single common ancestor mentioned carries the fimH30 allele (181).The study found that 91% of the CTX-M-15-producing isolates were responsible for the formation of theH30-Rx sub-clone. Lim et al. indicate that the H30-Rx was derived from a single common ancestor within the H30-R sub-clone that has already been discussed (140).
The sub-clones result in the formation of a Russian-doll-like configuration (Tamang et al.14).The configuration is responsible for the production of a lineage where one is more extensively resistant than a preceding sub-clone.
The confinement of the CTX-M-15 element to the H30-Rx sub-clone has a corresponding reaction to the ST131 gnome. According to Kim et al., the CTX-M-15 element has promiscuity within the ST131 genome (1572). In the study, the H30-Rx isolates were identified by a copy of the element which is usually found on the chromosome. In a separate study, Pfeifer, Cullik, and Witte indicate that the CTX-M-15 element is located in a region referred to as the IncFIItype plasmid (372).That notwithstanding, there are exceptional cases when the element occurs in both the chromosome and a plasmid (Banerjee et al. 6387).
The component in the samples with a chromosomal CTX-M-15 is usually placed in multiple sections. The study by Banerjee et al. had the insertion done in five different locations (6386). According to Banerjee et al., it is noted that the only two isolates with identical chromosomal insertion sites were recovered from grown up siblings with similar epidemiological traits (6386). The two individuals suffered from UTIs of varying severity (Banerjee et al. 6386).
In addition, it was believed that they had the same strain of ST131 (Banerjee et al. 6386).From the study and with respect to the 22 CTX-M- 15-producing strains, the CTX-M-15 elements do not give an indication of any kind of SNPs. Under such circumstances, Foxman argues against the comparison of a phylogenetic signal with respect to the host strain phylogeny (650). It is plausible that the chromosomal and parts of the plasmid CTX-M-15 components were obtained in a unique way.
They were obtained horizontally by H30-RX on different cases (Foxman 650). In spite of this, existing facts suggest something different within H30-Rx. To this end, it is noted that the chromosomally encoded CTX-M-15 is obtained from recurrent intragenomic movements. The movement is noted in the plasmid-located Tn3-like ISEcp1–blaCTX-M-15-orf477 compound. The case is different in independent horizontal acquisition events.
In the results section, it was observed that, as the infection progressed, pathogenic micro-organisms encountered multiple evrironements. According to Corvecet al., such a scenario causes the organisms to adapt (190). The same allows the coloy to thrive. To this end, it is necessary for pathogenic bacteria, like E. coli, to synthesise virulence factors. that will allow them to colonise a hostile environment and to survive the host immune and non-immune defences.
As part of this study the analysis of the sequence data was carried out using VirulenceFinder 2.1 Server to identifcation the acquired virulence genes among the number of 75 isolates from single nursing home
The study by Banerjee et al. sought to find out the prevalence of ST131 (6385). The same was conducted with respect to two of t gene’s sublones which are H30 and H30-Rx. The study was based on a case-control of infections brought about by extended-spectrum-lactamase (ESBL) producing E. coli strains (Banerjee et al. 6385).
Banerjee et al. compared these groups based ona number of factors (6385). The factors included virulence genotypes, antimicrobial resistance, and ESBL type (Banerjee et al. 6385). In this particular study, 267 E. coli isolates were collected over a duration ranging between 2007 and 2010. Banerjee et al. indicate that 100 of the isolates were ESBL-positive while the remaining 167 tested ESBL-negative (6385). There was a case-control study, as required for such a research design. The current study was also carried out upon the satisfaction of the laid out criteria.
In the study by Banerjee et al., fifteen health care-associated isolates were collected in the duration of the study (6386). As required, PCR-based methods were used to define E. coli phylogenetic group. Banerjee et al. note that (A, B1, B2, and D), ST131 and its H30 sub-clone (1, 2, 5), major CTX-M groups, the presence of blaCTX-M-15, and extended virulence genotypes [8–10] (6390).
In addition, the H30-Rx sub-clone was detected through PCR of a given single-nucleotide polymorphism (SNP) (G723A) within the allantoin- encoding gene, ybbW (2). Furthermore, primersAPfor63 (5’-GGTTGCGGTCTGGGCA-3’) and APrev66 (5’-CAATATCCAGCACGTT CCAGGTG-3’), with a cycling routine of 95°C for 8 min, 31 cycles of 94°C for 20 s and 72°C for 40 s, and a final extension at 72°C (for 5’), yielded a 194-bp amplicon (Banerjee et al. p. 6390).
Based on the results provided, it is noted that the resistance rating was obtained from the agents. The case applied to specific samples found to be resistant. To this end, 49 ESBL-positive ST131 isolates had a prevalence of 98%. The number was a representation of the H30 sub-clone. On the other hand, 44 of the isolates were a representation of the H30-Rx sub-clone. The results were a sharp contrast to the 22 ESBL-negative ST131 isolates from the same study.
According to Banerjee et al., only 14 represented the H30 sub-clone (6386). The figure represented a total of 64% at a P_ 0.001 level of significance. In addition, only 3 reported cases of H30-Rx sub-clone. The figure was 34% of the total at P _ 0.001. Banerjee et al. noted that H30 ST131 sub-clone isolates were increasingly resistant to fluoroquinolone resistant (6386). The level of resistance stood at 98%. In addition, 77 percent of the isolates tested positive for ESBL positive. However, a small number of non-H30 ST131 isolates showed resistance (Banerjee et al. 6386).
The results from this study compare to other studies with respect to the resistance brought about by the two sub-clones mentioned. For instance, in the study by Hornsey, Phee, and Wareham, the resistance levels were recorded as high as 69% (5952). The implications of these results are such that the effects of the fluoroquinolone are brought into sharp focus.
The performance of the drugs is inhibited by the nature of the isolates mentioned. According to Carter et al., drugs used to combat E. coli ought to be continually structured to withstand the resistance brought about by the sub-clones (4228). To this end, constant research on the fundamentals of the resistance should be undertaken to improve upon their respective performance.
VirulenceFinder
Just like the ResFinder, the VirulenceFinder is a technique used to study the genome of bacterial species. The major difference between the two however is that ResFinder is used for the detection of antibiotic resistant genes while VirulenceFinder is used for the detection of those genes that confer virulent characteristics to an organism. Most of the pathogenic strains of E. coli have virulent genes or factors. In the study, there was need to carry out the test in order to find out which of the organisms acquired the virulent genes.
The acquisition of the virulent genes makes an organism pathogenic. In a worst case scenario, an organism may acquire the virulent and the antimicrobial resistant genes (Corvec et al. 189). Control and management of such a microbe would prove to be a difficult task. In this study, the VirulenceFinder technique was applied on the isolates that had already undergone the ResFinder test.
Among the virulent factors that were investigated include nfaE, Adherence protein (iha), serine protease autotransporters of enterobacteriaceae (sat) that is also commonly reffered to as SPATE, Increased serum survival (iss), P-related fimbriae regulatory gene (PrfB), Glutamate decarboxylase (gad), Long polar fimbriae (ipfA), SenB, as well as, Siderphore receptor (ireA).
Just like the case of ResFinder, the VirulenceFinder test had equally varying results. Some of the genes were more prevalence among the isolates than the others. The prevalence of nfaE factor was moderate. It was found to be 69 percent. The iha, sat, iss, and Prfb factors on the other had some of the highest VirulenceFinder prevalence recorded.
Their prevalence was at 93, 92, 97, and 98 percent respectively. The factors are therefore of more importance in the study of medicine since they pose the greatest risk to the population following the acquisition of the virulent factors. Some of the factors however recorded the lowest prevalence rates. The factors are gad, ipfA, senB, and ireA. The four factors had a prevalence rate of 6, 5, 1, and 1 percent respectively. These factors are not of much concern in health management since they are not easily transmittable from one isolate to another.
With reference to the tabulated results for the VirulenceFinder that were obtained from the same isolates that had been subjected to ResFinder.
There is a great variation in the VirulenceFinder results. The prevalence of the virulent factors is at the extreme ends, either too high or too low. The range of the results is 97 percent. The figure is obtained using a specified computation method. It involves subtracting the least value 1 percent from the highest value (Mora et al. 1).
In this case, the highest figure is 98 percent. The study by Mora et al. sought to examine virulence factors (1). In a preliminary study of virulence in murine sepsis model, the study made use of 4 ST131 isolates. The research undertaking found that all the isolates gave rise to 100% lethality. The same is an indicator of rapid and extensive emergence ofCTX-M-15-producing ST131 clonal group.
In cases where the clonal group emerges, research attributes the same to elevated degrees of antibiotic resistance coupled with a corresponding increase in the virulence-gene scores. That notwithstanding, the study by Lim et al. indicates the possibility of a non-uniform virulence with respect to the isolates(140).
The same is vividly illustrated in the current study. The study by Mora et al. indicates that 30% of the ST131 isolates had a corresponding lethality of 0%. It is noted that the varying nature of virulence prevalence in ST131 clinical samples is a common phenomenon in such studies. The same is an indicator that combating the E. coli requires an element of diversity.
Interestingly, it has been observed that in most cases, virulence patterns of specific virotypes have an extremely high level of lethality. Mora et al. indicate lethality rates that are as high as 80%. According to Mora et al., ST131 samples drawn from virotypes A, B, and C indicated identical results (7). They showed high prevalence of final lethality. The figure stood at 80% (Mora et al. 8). The findings are significant to similar studies in this field, including the current research undertaking.
The results indicate the crux of the study in that the sub-clones of the E. coli genes pose a great health hazard to organism. On the contrary, in cases where there is a variable virulence pattern, the lethality is extremely low. Banerjee et al. report that in such cases, only lesions emerge (6387). The phenomenon of varied lethality is referred to as in vivo virulence diversity. Mora et al. argue that the ST131 clonal group is significantly related to genes (6).
To this end, the group is an important indicator of the genetic variability of materials (Mora et al. 6).The situation is brought about by a number of factors. One of them is the molecular typing of the housekeeping genes.
Most of the said genes are responsible for the emergence of the PSTs. According to Mora et al., the PSTs are part of the fimH subtyping that helps to define various clonotypes (6). The study also indicates PFGE profiles which are sourced from the XbaI digested DNA normally. Mora et al. explain the incidence in their study (6).
Mora et al. noted that O25b:H4samples of virotypes A, B, and C displayed identical ST [PST43] (6). In addition, the scholars found that similar fimH30 subtype and their PFGE trends were brought together in dendrogram. The identical ratio stood at 75.8% (Mora et al. 6). Mora et al. continue to argue that O25b:H4 isolates of virotype D were found in 3 varying STs (6). The STs were PST9, PST43, and PST527 (Mora et al. 6).
With respect to the assessment of virulence of ST131 few studies have focused on the isolates of serotype O16:H5. Mora et al. pioneered on the same and indicated thatisolate has a low virulence-gene score (9). In such cases did lethality with respect to virulence is very high. The same is indicated in the results reported by Mora et al. (7).
The research of Mora et al. showed that the E. coli O25b:H4 isolate from the virotypes A, B, and C causes a slow death. On its part, O16:H5 ST131 isolates recorded high levels of virulence. The results were in line with reported clinical severity in similar cases (Mora et al. p. 7).
It is noted that isolateO16 ST131 is a leading factor behind pyelonephritis among both male and female patients. It is especially common during the reproductive age. The virulence results from Mora et al. suggest that clonotype H41 is closely relatedwith serotype O16:H5 (7). According to Mora et al. (7), the two are the major causes of the condition (Mora et al. 7). In the study, the importance of clonotyping, regarding the discrimination of subline ages of the clones is highlighted. There are specific clonotypes characterised with antimicrobial susceptibility properties that contribute to the virulence indicated in the current study.
The findings made by Mora et al. have a significant impact on the prevalence factors outlined in the current study (7). For example, Mora et al. note that 4 fimH types were related closely to serotype and virotype (7).
The clonotype mentioned in the current study is responsible for urinary tract infection. Similar results were found in the study carried out by Mora et al. where the same was the cause of the high lethality recorded (7).
It is noted that a number of epidemiological and clinical trends are closely linked to the virotype mentioned (Lautenbach et al. 1170). According to Lautenbach et al., the characteristics help to explain albeit partially or the various patterns of virulence that are exhibited by ST131 (1165). The virulence of three ST131 samples was analysed by Mora et al. (7). The results were identical to five non-ST131 alternative nematodes. The five were derived from Caenorhabditis elegans and zebrafish embryos (Mora et al. 5).
The findings of low virulence for ST13isolates are brought about by a number of reasons. Banerjee et al. enumerate some of the factors behind these cases (6388). According to Banerjee et al., the lack of a number of virulence factors is a possible explanation of the situation (6388). To this end, the scholars cite HlyA and Cnf1as some of the significant elements behind the occurrence (Banerjee et al. 6388). The same applies from the results reported in the current study.
There are other results which indicate virulence. The high genetic variability within the ST131 clonal group has a direct impact on the lethality of the isolates selected. For study purposes, rodents and canines provide suitable results. Clonotyping also helps to realised the virulence results intended for a study.
Conclusion
The E. coli ST131 is an ESBL producing group of micro organisms. For this reason, they are associated with resistance to antimicrobial agents. Many scientists across the globe have embarked on studying aspects associated with these microbes. In the current study, isolates were collected from a nursing home and the Khartoum hospital.
Several tests were carried out to determine the various characteristics associated with the microbes. One of the most important tests carried out is phenotypic ESBL confirmation. The aim of the test was to determine which of the isolates produced ESBLs and the type of this enzyme that they produced. The test is important since it helps determine what antimicrobial agents the isolate is resistant to.
The isolates were also subjected to H30-RX PCR. The main aim of the test was to determine the prevalence of ST131 H30-RX sub-clones among the isolates. Of the 88 isolates tested, 70 percent were H30-RX PCR positive showing that there was a higher prevalence of the sub-clone. For the H30 PCR, 60 percent of the isolates tested positive.
The results clearly show that H30-RX sub-clone is more prevalent compared to the H30. Scientific studies have determined that the reason behind this is that H30-RX sub-clones account for the over 91 percent of the CTX-M-15 producing ST131 E. coli. The enzyme enables the microbes to be resistant to antimicrobial agents, especially fluoroquinolones.
For this reason, they continue to persist in the gut and the urinary tract of their host even after the administration of antibiotics. Once the normal microflora balance is upset following the destruction of harmless bacteria, diseases set in causing serious health problems, such as the urinary tract infection (UTI). Today, ST131 H30-RX is the main group of E. coli that causes urinary tract infections.
The isolates were also examined on their ability to acquire antibiotic resistance genes. The study showed that indeed the ST131 have in the past acquired these genes. Different genes are responsible for resistance to different antimicrobial agents. At the present, genes responsible for resistance to Beta-lactam, chloramphenicols, trimethoprim, tetracycline, sulphonamide, aminoglycosides, and fluoroquinolones have been identified. The genes were also found to be present in the isolates under investigation.
The results of the study were therefore a testimony to the fact that ST131 clones have the ability to acquire and transfer antibiotic resistance genes. Scientists have also linked the characteristic to their widespread nature. Once they infect the host, they are able to corrupt the genome of other previously harmless microbes.
The isolates were also tested for virulent factors. Bacterial species are known to acquire virulent factors with the aim of helping them survive even the harshest environmental conditions. The ST131 H30-RX sub-clones are known to be the most virulent group of microbes in the species. As a result, they have been in a position to multiply even upon the administration of a wide range of antimicrobial agents. Following the acquisition of these factors, the microbes are also in a position to resist natural defence by the body of the host.
Research has also shown that the microbes are in a position to confer the virulent factors to other microbes. As a result, more microbes are acquiring resistance to antimicrobial agents. The microbes are also in a position to adapt easily to harsh conditions subjected to the microbes by the host’s body.
Works Cited
Banerjee, Ritu, Ari Robisek, Michael Kusikowski, Stephen Porter and Brian Johnston. “Molecular Epidemiology of Escherichia Coli Sequence Type 131 and its H30 And H30-Rx Sub-clones among Extended-Spectrum-Β-Lactamasepositive and -Negative E. coli Clinical Isolates from the Chicago Region, 2007 To 2010.” Antimicrobial Agents and Chemotherapy 57.12 (2013): 6385-6388. Print.
Carter, Michael, Karen Oakton, Marina Warner and David Livermore. “Detection of Extended-Spectrum β-Lactamases in Klebsiellae with the Oxoid Combination Disk Method.” Journal of Clinical Microbiology 38.11 (2000): 4228-4232. Print.
Clermont, Olivier, Neil Woodford, David Boyd, Andrew Fox, Tarek Gibreel, Mathew Upton, Hiran Dhanji, Erick Denamur, Catherine Branger, Guillaume Arlet, Sophie Vimont, Thierry Frank, Jean Sarthou, Etienne Ruppe, Patrice Nordmann, and Michael Mulvey. “Rapid detection of the O25b-ST131 clone of Escherichia coli encompassing the CTX-M-15-producing strains.” Journal of Antimicrobial Chemotherapy 64.2 (2009): 274-277. Print.
Corvec, Stephane, Nathalie Caroff, Eric Espaze, Julie Marraillac, Henri Drugeon and Alain Reynaud. “Comparison of Two RT-PCR Methods for Quantifying ampC Specific Transcripts in Escherichia Coli Strains.” FEMS Microbiology Letters 228.2 (2003):187-191. Print.
DeVincent, Stephen, and Richard Reid-Smith. “Stakeholder Position Paper: Companion Animal Veterinarian.” Preventive Veterinary Medicine 73.2-3 (2006): 181-189. Print.
Foxman, Betsy. “Extended-Spectrum β-Lactamase–Producing Escherichia Coli in the United States: Time to Rethink Empirical Treatment for Suspected E. coli Infections?” Clinical Infectious Diseases56.5 (2012): 649-651. Print.
Hornsey, Michael, Lynette Phee, and David Wareham. “A Novel Variant, NDM-5, of the New Delhi Metallo-β-Lactamase in a Multidrug-Resistant Escherichia Coli ST648 Isolate Recovered from a Patient in the United Kingdom.” Antimicrobial Agents and Chemotherapy 55.12 (2011): 5952. Print.
Kim, J, Y Lim, Y Jeong and S Seol. “Occurrence of CTX-M-3, CTX-M-15, CTX-M-14, and CTX-M-9 Extended-Spectrum ß-lactamases in Enterobacteriaceae Clinical Isolates in Korea.”Antimicrobial Agents Chemotherapy 49.4 (2005):1572-1575. Print.
Lautenbach, Ebbing, J Patel, WBilker, P Edelstein and N Fishman. “Extended-Spectrum  -Lactamase-Producing Escherichia coli and Klebsiella pneumoniae: Risk Factors for Infection and Impact of Resistance on Outcomes.” Clinical Infectious Diseases 32.8 (2001): 1162-1171. Print.
Lim, Suk-Kyung, Hee-Soo Lee, Hyang-Mi Nam, Suk-Chan Jung and You-Chan Bae. “CTX-M–type Betalactamase in Escherichia Coli Isolated from Sick Animals in Korea.”Microbial Drug Resistance 15.2 (2009):139-142. Print.
M’Zali, Fatima, Aroonwadee Chanawong, Kevin Kerr, David Birkenhead and Peter Hawkey. “Detection of Extended-Spectrum Blactamases in Members of the Family Enterobacteriaceae: Comparison of the Mast DD Test, the Double Disc and the Etest ESBL.”Journal of Antimicrobial Chemotherapy 45, (2000): 881-885. Print.
Mora, Azucena, Ghizlane Dahbi, Cecilia Lopez, Rosalia Mamani, Juan Marzoa, Sara Dion, Betrand Picard, Miguel Blanco, Maria Alonso and Jorge Blanco. “Virulence Patterns in a Murine Sepsis Model of ST131 Escherichia Coli Clinical Isolates Belonging to Serotypes O25b:H4 and O16:H5 are Associated to Specific Virotypes.” PLoS ONE 9.1 (2014): 1-9. Print.
Nicolas-Chanoine, Marie-Helene, Xavier Bertrand, and Jean-Yves Madec. ” Escherichia coli ST131, an Intriguing Clonal Group.” Clinical Microbiology Reviews 27.3 (2014): 543-574. Print.
Paterson, David, Robert Bonomon. “Extended-Spectrum B-Lactamases: A Clinical Update.” Clinical Microbiology Reviews 18.4 (2005): 657-686. Print.
Perez-Perez, Javier, and Nancy Hanson. “Detection of Plasmid-Mediated ampC Beta-Lactamase Genes in Clinical Isolates by using Multiplex PCR.” Journal of Clinical Microbiology 40.6 (2002): 2153-2162. Print.
Pfeifer, Yvonne, Angela Cullik, and Wolfgang Witte. “Resistance to Cephalosporins and Carbapenems in Gram-Negative Bacterial Pathogens.” International Journal of Medical Microbiology 300.6 (2010): 371-379. Print.
Tamang, Migma, Hyang-Mi Nam, Su-Ran Kim, Myung Chae, Geum-Cab Jang, Suk-Chan Jung and Suk-Kyung Lim. “Prevalence and Molecular Characterisation of CTX-M b-Lactamase–Producing Escherichia coli Isolated from Healthy Swine and Cattle.”Foodborne Pathogens and Disease 10.1(2013): 13-20. Print.
Yokota, Shin-Ichi, Toyotaka Sato, Torahiko Okubo, Yasuo Ohkoshi, Tamaki Okabayashi, Osamu Kuwahara, Yutaka Tamura, and Nobuhiro Fujii. “Prevalence of Fluoroquinolone-Resistant Escherichia coli O25:H4-ST131 (CTX-M-15-Nonproducing) Strains Isolated in Japan.” Chemotherapy 58.1 (2012): 52-59. Print.