Lesson 4: Acquisition and spread of antibiotic resistance PDF

Lesson 4: Acquisition and spread of antibiotic resistance Prepared by Dr/ Essam KOTB Associate Professor of Microbiology, IAU...

Lesson 4: Acquisition and spread of antibiotic resistance Prepared by Dr/ Essam KOTB Associate Professor of Microbiology, IAU [email protected] +2/01121343400 +966/0563533550  Acquired resistance results from mutation or horizontal gene transfer. This explains why antibiotic resistance arises and spreads so rapidly.  a) Acquired resistance by of mutations – The best example is Mycobacterium tuberculosis (the causative agent of TB) where resistance to all therapeutic agents is caused by mutation. – How does mutation of a bacteria’s DNA result in antibiotic resistance? Genetic informations, encoded by DNA, are converted into proteins which are required for the structure and function of bacteria. Antibiotics often exert their effects by binding to proteins that are crucial to the structure or function of the bacterial cell. Structural changes to an antibiotic target protein by mutation prevent the antibiotic from binding i.e make the target resistant to Figure 2 Genetic mutations can alter the amino acid sequence of a protein. Transmission of mutations by vertical gene transfer As mutations are permanent changes in the DNA sequence that makes up a gene, the mutated bacterium will be resistant to the particular antibiotic for infinity. During binary fission, the mutated chromosomal DNA is copied, so that each new daughter cell inherits an exact copy of the parent cell’s chromosomes. Therefore, these resistant mutations are transferred from a parent cell to its offspring which is called vertical gene transfer. Fig. The stages of vertical gene  b) Acquired resistance by horizontal gene transfer Horizontal gene transfer is the mechanism of spread of antibiotic resistance genes between unrelated bacteria. This is particularly concerning because it can result in multidrug-resistant bacterial strains (superbugs) such as MRSA. Unlike vertical gene transmission, where chromosomal DNA is replicated and then transferred from parent cells to daughter cells through binary fission, mainly plasmids or transposons are transferred by horizontal gene transfer. This is the process of swapping genetic information between two unrelated cells does not require binary fission and can occur between bacteria of the same generation, not just between parents and daughters (Figure). Fig. The differences between horizontal and vertical gene transmission.  From the previous, one can conclude why horizontal gene transfer is the primary mechanism of spreading antibiotic resistance? – Horizontal gene transfer allows plasmids and transposons carrying antibiotic resistance genes to spread rapidly between different bacteria. – Therefore, species of bacteria that are intrinsically sensitive to a given antibiotic rapidly acquire resistance genes, making them resistant to treatment with that antibiotic. Mechanisms of horizontal gene transfer: a- Conjugation It is a process in which plasmids are transferred between two bacteria via a conjugation tube and a sex pilus. Since antibiotic resistance genes are often located on transposons and plasmids, a copy of drug resistance plasmid called R plasmid can transfer from one bacterium to another. Fig. The process of conjugation. (a) A hollow pilus connects two bacteria and plasmid DNA is b- Transformation It is a process in which bacteria can take up pieces of DNA across cell membrane from their environment from a lysed resistant bacterium. This DNA incorporated into the genome of the recipient bacterium conferring antibiotic resistance. Transformation occurs naturally between some bacteria, such as Streptococcus pneumoniae and Haemophilus influenza. Fig. A bacterium taking up DNA from the environment by transformation. c- Transduction It is the process in which the transfer of DNA from resistant bacteria to sensitive bacteria is mediated by a temperate bacteriophage. When bacteriophages infect a bacterial cell, they insert their DNA into the bacterial cell genome. When it is time for the virus to replicate, it excises its DNA from the bacterial genome. However, during excision some of bacterial DNA is accidentally taken and incorporated into the newly made virus. When these newly made viruses infect different bacterial species, they carry this bacterial DNA, which contain antibiotic resistance genes, and insert it into the genome of the new host bacterium. Fig. Process of transduction. When bacteriophage DNA, shown by a black dotted line, is excised from the bacterial genome it carries with it some bacterial DNA, shown in blue, from the infected bacteria. This DNA is incorporated into new bacteriophage particles which are released and infect new bacteria of a different species. The bacterial DNA from the original bacteria, in blue, is incorporated into the genome of the newly infected bacteria.  Case study: resistance to cephalosporins  In lesson’s 3 case study, you looked at the molecular mechanisms of resistance to cephalosporins and were introduced to ESBLs. The most common class of ESBLs in Europe is the CTX-M-type ESBL. Now, you will explore how bacteria acquire resistance to cephalosporins through horizontal gene transfer and the mutation of CTX-M-type ESBLs. You will begin by looking at how the presence of CTX-M- type ESBLs has changed in the UK over recent years. First, look at Figure 11 which shows the percentage of E. coli isolate producing CTX-M-type, or other, ESBLs between 2002 and 2016. Figure 11 UK susceptibility survey data for  Using the data in Figure 11, how has the proportion of isolates producing CTX-M-type ESBLs changed over time? CTX-M-type ESBLs emerged in the late 1990s and were first reported in the UK in 2002. Suggest one possible reason for the change in frequency of these ESBLs in the UK over time?.  In Activity 7 in lesson 1, you looked at how resistance to cephalosporins had changed in the UK over time (Figure 12). By comparing the data in Figures 11 and 12 do you think that the occurrence of CTX-M-type ESBLs is a good indicator of the rate of resistance to cephalosporins? What challenges might these changes in the prevalence of CTX- M-type ESBLs present to health care? Figure 12 Resistance to cephalosporins in the UK  The origin of CTX-M-type ESBLs Unlike most acquired β-lactamases, for which the source remains unknown, the source of CTX-M genes has been identified in genus Kluyvera. Kluyvera are soil bacteria which are associated with plant roots and are non-pathogenic to humans. The resistance of Kluyvera to cephalosporin generations is an example of what type of resistance? – It is an example of intrinsic resistance. These chromosomal CTX-M genes have been captured from the chromosomal DNA of Kluyvera by horizontal gene transfer and incorporated into plasmids of other bacterial types, including E. coli and Klebsiella pneumoniae (Figure 13). The transfer occurs in the human gut and in the environment and is fundamental to their global spread. Therefore, plasmids Figure 13 Plasmid map of a CTX-M- containing plasmid isolated from E. coli. The CTX-M gene is shown in  Plasmids carrying CTX-M genes often carry bacteriophage- related sequences or genes that are required for the formation of pili. – The presence of bacteriophage-related sequences in some CTX-M-containing plasmids suggests horizontal gene transfer by transduction. – The presence of genes required for the formation of pili suggests horizontal gene transfer via conjugation which requires a pilus linking the donor and recipient bacteria. The most concerning feature of CTX-M-containing plasmids is their ability to acquire additional antibiotic resistance genes. If they acquire resistance genes to carbapenems, which are frequently used to treat cephalosporin-resistant infections, it could seriously challenge the treatment of infections. Mutations in CTX-M CTX-M preferentially acts on certain cephalosporins: – Cefotaxime is easily recognised and inactivated by CTX-M. – While the bulkier cephalosporin ceftazidime is poorly recognised by CTX-M. As a consequence, infections caused by bacteria that produce CTX-M can be treated with ceftazidime. This specificity is based on the structure of the CTX-M β-lactam binding site (blue arrow) which only allows the efficient recognition of penicillins and cefotaxime. Amino acid mutation at location of the blue star generates the ceftazidimase activity of CTX-M. Fig. The shape of a CTX-M-type ESBL protein.  For bacterial adaptation, the specificity of CTX-M can be modified by point mutations which improve the specificity of CTX-M for ceftazidime. Altering this amino acid allows the bulkier ceftazidime to be more easily accommodated in the β-lactam binding site. As a result, infections caused by bacteria producing this mutated version of the CTX-M are not treatable with ceftazidime. These CTX-M variants have been isolated from clinical strains of E. coli and has been selected due to the increasing use of ceftazidime.