Information

What is the specific use of a capsule in E.coli?


In evolutionary terms, will the capsule cease to exist someday or will it improve ? Does it provide any boost to the organism in any way which harms us ? Can't we remove capsules or engineer E.coli without capsule ?


According to wikipedia, the capsule contains tightly packed polysaccharides that preform several functions including keeping bacteriophages out, blocking recognition by macrophages, and holding water during dessication. Given the wide diversity of bacteria with capsules, it's probably been around for a long time, which implies it isn't going away any time soon.

If the capsule prevents the E. coli from being taken up by macrophages, then yes, it could allow the bacteria to harm us more effectively. This is one form of a virulence factor.

With proper conditions we probably could develop a bacteria with little to no capsule. We have developed L-form bacteria, which don't have peptidoglycan cell walls. It would seem to me that the cell wall is more important than the capsule, but L-form bacteria are very fragile. I don't know what advantage would be gained by producing capsule free bacteria. There would probably be more to gain from making bacteria with different capsules, perhaps carrying proteins that recognize certain substances and produce a signal, allowing the bacteria to be used as a biosensor, or to accumulate toxic compounds for bioremediation.

references: http://en.wikipedia.org/wiki/Bacterial_capsule http://en.wikipedia.org/wiki/L-form_bacteria


Capsule and lipopolysaccharide

Pathogenic E. coli require many different virulence factors which allow them to invade the host, evade host immune defenses and colonize specific niches in the host where they can cause disease. The first interactions between E. coli and its host occur at the outer membrane and are mediated by proteins and carbohydrate-containing macromolecules (glycoconjugates) on the bacterial and host cell surfaces (Figure 17.1). Bacterial glycoconjugates provide crucial defenses against different elements of the innate and acquired immune system, as well as generating tremendous diversity in surface antigenicity. In most Gram-negative bacteria, the outer membrane is composed predominantly of the glycolipid known as lipopolysaccharide (LPS). In E. coli , this complex molecule is composed of three structurally distinct regions the hydrophobic anchor called lipid A, a core oligosaccharide, and the long-chain polysaccharide called O antigen (or O-polysaccharide O-PS). The differences in E. coli O-PS structures give rise to more than 180 distinct O antigens and these have been exploited in serotyping classification methods (Orskov et al., 1977). Many E. coli isolates also produce another long-chain polysaccharide, known as capsular polysaccharide (CPS). CPS provides another major surface antigen, called the K-antigen, named after the German term ‘kapsel’. There are more than 80 different K antigens in E. coli . The means of attachment of CPS to the cell surface is not known in all cases but these polymers create a coherent structural entity (the capsule) that is visible by light microscopy and extends 50–100 nm from the cell surface. As a result, the capsule often masks underlying O antigens in serotyping studies that exploit specific antisera and whole-cell agglutination methods. A single E. coli isolate can produce one O- and one K-antigen.

Together, LPS and CPS represent major virulence factors, which have been the target of numerous vaccines and therapies, and they provide the focus of this chapter. However, they are not the only glycoconjugates produced by E. coli . All isolates produce a polysaccharide called enterobacterial common antigen that provides resistance to organic acids in E. coli (Barua et al., 2002) and to bile salts and detergents in Salmonella (Ramos-Morales et al., 2003). Under certain stress conditions (e.g. perturbations of cell envelope integrity), some E. coli isolates also produce an exopolysaccharide called colanic acid which has a biosynthetic pathway that is part of the Rcs-regulon (Majdalani and Gottesman, 2005). Unlike CPS, colanic acid is poorly retained at the cell surface. Like several other bacterial species, the biofilm mode of growth in E. coli is supported by formation of bacterial cellulose and a by-polymer of N -acetylglucosamine (PNAG) (Cerca and Jefferson, 2008 Saldana et al., 2009). However, some commensal E. coli isolates also produce a polysaccharide of unknown structure that impairs biofilm formation by other bacteria including Staphylococcus aureus , potentially affecting community dynamics (Rendueles et al., 2011). Similar anti-adhesion properties have been reported for certain E. coli CPSs (Valle et al., 2006). In summary, the cell surfaces of E. coli isolates are rich in complex carbohydrates and these molecules play diverse, niche-dependent roles in the physiology of E. coli . In this chapter, we will focus only on LPS and CPS.

Structure and biosynthesis of E. coli LPS

The LPS molecule contains three regions, the lipid A, core, and O-PS. Lipid A is the most highly conserved portion of the molecule. The typical structure contains two phosphorylated glucosamine residues with six acyl chains (Figure 17.2A), though this can be modified in several places to alter the biological properties of the molecule (reviewed in Raetz et al., 2007) (see below). The core oligosaccharide can be divided into two regions. The inner core consists of two residues of 3-deoxy- D – manno -oct-2-ulosonic acid (Kdo) and three residues of L -glycero- D – manno -heptose (Hep) but it may also be modified with a number of other groups such as phosphate, pyrophosphorylethanolamine, or additional sugars (Figure 17.2B). This region is highly conserved in E. coli and Salmonella and is important for membrane stability (reviewed in Heinrichs et al., 1998). For example, loss of the phosphate moieties on Hep residues compromises outer membrane integrity and, in Salmonella , attenuates virulence (Yethon et al., 2000). The outer core serves as the point of attachment for O-PS and contains sugars such as glucose (Glc), galactose (Gal), glucosamine (GlcN), and N-acetylglucosamine (GlcNAc). Despite the potential for great diversity, there are only five known core types in E. coli , K-12, R1, R2, R3, and R4 (Heinrichs et al., 1998 Kaniuk et al., 2004). The core type R1 is most commonly found in E. coli causing extraintestinal infection while the verotoxin-producing E. coli isolates are predominantly R3 (Figure 17.2B).

The O antigen is highly variable and typically consists of repeat units of 2–5 sugars in a polymer that can be more than 100 sugars long. The range of repeat unit numbers is specific for a particular strain and is controlled by different strategies, depending on which of the two biosynthetic pathways is used for O-antigen biosynthesis: the Wzy-dependent pathway or the ABC transporter-dependent pathway (Figure 17.3) (Raetz and Whitfield, 2002). In all cases, biosynthesis of O antigens begins at the cytoplasmic face of the inner membrane using the lipid carrier undecaprenyl phosphate. In E. coli, the enzyme WecA initiates O-PS biosynthesis by transferring GlcNAc-1-phosphate from UDP-GlcNAc to undecaprenyl phosphate to form undecaprenyl pyrophosphoryl-GlcNAc. In some cases, this is epimerized to undecaprenyl pyrophosphoryl-GalNAc (Rush et al., 2010). The GlcNAc/GalNAc residues occur in each repeat unit in O-PSs formed by the Wzy-dependent pathway, or just once at the reducing terminus in the ABC transporter-dependent process.

In the Wzy-dependent pathway, individual repeat units are synthesized on undecaprenyl pyrophosphoryl-GlcNAc/GalNAc using nucleoside phosphate sugar donors by enzymes encoded in the locus. These repeat units are then flipped to the periplasmic side of the inner membrane by the flippase Wzx and polymerized into the complete O antigen by the polymerase Wzy. Wzy transfers the growing glycan from one undecaprenyl pyrophosphate carrier to the incoming lipid-linked repeat unit. Insight into this pathway has benefitted from model systems, including the O7 and O86 antigens (Table 17.1), studied in the laboratories of Miguel Valvano and George Peng Wang, respectively. In the ABC transporter-dependent pathway, the O antigen is synthesized entirely at the cytoplasmic face of the inner membrane through the sequential action of glycosyltransferases that add sugars to undecaprenyl pyrophosphoryl-GlcNAc, before the completed chain is exported via an ABC transporter. This process is less common than the Wzy-dependent pathway and the resulting repeat-unit structures tend to be less elaborate. The serotype O8/O9/O9a antigens provide influential prototypes for this type of assembly process and many of the steps have been resolved by the research group of Klaus Jann and the Whitfield lab (Table 17.1). The E. coli WaaL ligase can operate effectively with nascent undecaprenyl-linked O-PSs from either pathway.

The LPS species extracted from a bacterial cell shows heterogeneity best illustrated by profiles separated by SDS-PAGE (Hitchcock and Brown, 1983). The most obvious variations occur in the chain-lengths of O-PS, evident as a ladder of high-molecular-weight molecules. The length of the O-PS has important functional consequences (see below) and is established by different mechanisms, depending on the assembly pathway. In the Wzy-dependent pathway, the length of the O antigen is controlled by the protein Wzz, a transmembrane protein with a large periplasmic domain that belongs to the polysaccharide co-polymerase (PCP) family (reviewed in Cuthbertson et al., 2009 Morona et al., 2009). In the ABC transporter-dependent pathway, chain length of some O-PS molecules is controlled by the addition of novel residues (ones not found in the repeat-unit domain) to the non-reducing end of the glycan (reviewed in Cuthbertson et al., 2010). These residues may be alternative sugars or non-glycose moieties and they not only prevent further elongation of the glycan, but are also required to engage the ABC transporter for export across the inner membrane. However, other O antigens in this pathway engage the ABC transporter in the absence of any identifiable terminating group and coordination of the activities of the O-PS elongating enzymes and the ABC transporter determine O-PS chain length (Cuthbertson et al., 2010).

Most LPS modifications occur at the periplasmic face of either the inner or outer membrane and, as such, can serve as markers indicating the stage of transport. These include modification of the phosphate groups with 4-amino-4-deoxy-L-arabinose (L-Ara4N) or ethanolamine by enzymes found in the inner membrane, which increases bacterial resistance to innate immune defenses (see below). The outer membrane protein PagP adds a secondary palmitate in an acyloxyacyl linkage to the hydroxy-myristate located at the 2-position. The resulting heptaacyl species is a much less potent activator of cytokine induction (Raetz et al., 2007). PagP is normally latent in E. coli but is activated by membrane perturbations and by defects in acylation of the 3´ position of the diglucosamine backbone with a myristol residue. However, this modification can alter host–pathogen interactions in other ways, because PagP activity in E. coli O157 has an indirect effect on the completion of the LPS core oligosaccharide, leading to loss of O antigen and serum sensitivity (Smith et al., 2008).

Structure and biosynthesis of E. coli CPSs

The CPSs of E. coli have been subdivided into four groups based on structural and genetic criteria (reviewed in Whitfield, 2006). Group 1 and 4 capsules share the same mode of synthesis, a Wzy-dependent pathway, and mainly differ in the chromosomal locations of key genes. Similarly, groups 2 and 3 capsules are both produced by ABC transporter-dependent processes but the genes are organized differently within the locus.

Group 1 (and 4) CPS are found in E. coli isolates causing intestinal infections. They are heteropolymers of repeating sugar units, as in many O antigens. Biosynthesis uses a process identical to the Wzy-dependent O antigens but the pathways diverge once the polymerized glycan is formed at the periplasmic face of the inner membrane (Figure 17.4). O antigens enter the LPS assembly pathway by ligation to lipid A-core and are transferred to the surface via Lpt proteins. Capsular K antigens have their own surface assembly process that involves four key components: (i) Wzc, an inner membrane PCP protein belonging to the PCP-2a subfamily (ii) Wzb, a cytoplasmic protein tyrosine phosphatase (iii) Wza, an outer membrane lipoprotein belonging to the OPX (outer membrane polysaccharide export) protein family (Cuthbertson et al., 2009) and (iv) an accessory outer membrane protein called Wzi. Understanding the group 1 CPS translocation processes has come from studies with the serotype K30 prototype in the Whitfield laboratory. In O-antigen biosynthesis, the corresponding PCP protein belongs to sub-family 1 and its action appears to be confined to regulating the polymerization activity, although the exact mechanism is still unknown. PCP-2a proteins are more complex. Wzc certainly affects polymerization but this activity (and capsule formation) is dependent on the activity of a C-terminal tyrosine autokinase domain, as well as the dephosphorylation of these residues by the Wzb phosphatase (Wugeditsch et al., 2001). The autokinase domain is absent in PCP-1 family members. The additional role played by Wzc is mediated by its periplasmic domain, which is larger in PCP-2a proteins, and interacts with the extensive periplasmic region of Wza (Collins et al., 2007). The crystal structure of Wza reveals an octomer which forms a large periplasmic barrel connected to an outer membrane channel (Dong et al., 2006). The outer-membrane pore is formed by one α-helix contributed by each protamer and was the first example of an outer-membrane channel which is not a β-barrel (Dong et al., 2006). It is not known how the Wzc/Wza complex transports the capsule to the outer membrane or what the nature of the reducing end of the exported polymer is i.e. is the CPS attached to an anchoring protein or lipid. In O-antigen biosynthesis, the ligase (WaaL) releases the nascent glycan from the undecaprenyl pyrophosphate carrier and links it to lipid A-core. There is no corresponding activity in the biosynthesis of group 1 and 4 K antigens. The CPS could retain its linkage to undecaprenyl pyrophosphate, or it could potentially be released by a leaky polymerization step, where the transfer of the growing chain to the incoming repeat unit may be incomplete. Wzi is important for surface retention of group 1 CPS although its precise function is not yet known and it is not present in group 4 systems (Rahn et al., 2003). The Wza-Wzb-Wzc proteins are highly conserved in all isolates possessing a group 1 or 4 capsule, indicating that they do not recognize any particular glycan repeat-unit structure. Group 1 K antigens are typically co-expressed with an O antigen synthesized by the ABC transporter pathway (e.g. O8/O9/O9a) and the corresponding gene clusters are both located near his . The group 1 gene cluster is allelic with genes for colanic acid biosynthesis, so expression of a capsule and colanic acid are mutually exclusive. However, group 4 K antigens really emphasize the parallels between O- and K-antigen biosynthesis. In these cases, the structures of the repeat units are identical and are produced by the same chromosomal locus. Some undecaprenyl pyrophosphate-linked molecules are diverted by WaaL into LPS, while others enter a CPS translocation pathway by Wza-Wzb-Wzc proteins that are encoded by a separated locus elsewhere on the chromosome (Peleg et al., 2005). Serotype O111 provided the first example, and the use of the same repeat structure led to the early description of ‘ O-antigen capsules ’ (now group 4) (Goldman et al., 1982). It is now known that serotypes O127 and O157 also have group 4 capsules (Table 17.1) (Peleg et al., 2005 Shifrin et al., 2008). Isolates that can produce group 4 CPSs (unlike their group 1 counterparts) retain the genes for colanic acid production.

K antigens from extraintestinal pathogenic E. coli mostly fall into groups 2 and 3 (Whitfield, 2006). These include capsules from isolates causing urinary tract infections and meningitis. The production of many of these K antigens is temperature-regulated, unlike group 1 and 4 K antigens, with expression being ‘on’ at 37°C but ‘off’ at temperatures below 20°C. Some of the CPS structures resemble eukaryotic glycans and this is thought to aid virulence by preventing an effective immune response. For example, the K4 glycan is fructosylated chondroitin and the K5 glycan is heparosan, both similar to human glycosaminoglycans (Table 17.1). As well, the meningitis-causing K1 isolates possess a CPS containing α2,8-linked polysialic acid, which is identical in structure to the glycan found on neural cell adhesion molecule (NCAM) in the human brain. The K1 and K5 systems have been influential models for understanding the mechanisms of biosynthesis through studies in the laboratories of Eric Vimr, Willie Vann, and Ian Roberts. ABC transporter-dependent capsules are synthesized entirely on the cytoplasmic face of the inner membrane, similar to ABC transporter-dependent O antigens (Figure 17.4) (Whitfield, 2006). However, one critical difference is that undecaprenyl phosphate is apparently not involved (Finke et al., 1991). It has been shown that the mature capsular polysaccharide is attached at the reducing end to a phospholipid although the precise chemical structure has not been resolved (Gotschlich et al., 1981). In the essentially identical process in N. meningitidis , lipidation occurs before transport of polysialic acid CPS to the cell surface, but it is not known whether the glycan is synthesized directly on this phospholipid or if it is synthesized on another molecule before transfer to the phospholipid (Tzeng et al., 2005). Once the capsule is synthesized, it is transported through the inner membrane by the ABC transporter (KpsMT) (reviewed in Vimr and Steenbergen, 2009). The final steps of glycan translocation are mediated by KpsE, a representative of the PCP-3 subfamily, and KpsD, an OPX protein (Cuthbertson et al., 2009). A multiprotein export complex comprising KpsMTED is thought to span the cell envelope (Rigg et al., 1998 McNulty et al., 2006). Like the group 1 and 4 translocation export proteins, they are conserved in all isolates possessing a group 2 or 3 capsule, indicating that they also do not recognize any particular glycan repeat-unit structure. The genes encoding group 2 and 3 K antigens are located on the chromosome near serA and these glycans can be found together with a wide range of O serotypes, many formed by a Wzy-dependent system.

Evasion of host cell defenses

Mammals have evolved with the pressure of bacteria, viruses, and fungi and so have developed ways of dealing with microbial infections the most extensive defense machinery is the immune system. The human immune system consists of two branches, innate and adaptive immunity, between which there is extensive crosstalk. Innate immunity has developed as a first line of defense against challenges not necessarily seen before to protect the body from pathogens. Toll-like receptors (TLR) are the major effectors of innate immunity which recognize conserved structures in pathogens called pathogen-associated molecular patterns (PAMPs), somewhat of a misnomer, as they are found in non-pathogenic as well as pathogenic microbes in Gram-negative bacteria, these include LPS, CPS, flagella, and nucleic acids. Binding of TLRs to their respective ligands activates a signaling cascade, resulting in recruitment and activation of immune cells, which can then eliminate the threat. In addition to the cellular receptors, there are also soluble complement proteins found in serum, which can bind directly to bacterial cell surfaces leading to opsonization, phagocytosis, and formation of the membrane attack complex leading to lysis. LPS and CPS are important parts of the arsenal of virulence factors used by bacteria to circumvent these processes.

Lipid A structure influences susceptibility to polycationic peptides

Cationic antimicrobial peptides (CAMPs) are short peptides secreted by immune and epithelial cells in response to bacterial products, like LPS, and other inflammatory signals (reviewed in Brown and Hancock, 2006). It is well established that the structure of lipid A has a profound effect on the susceptibility of bacteria to CAMPs and polycationic drugs such as polymyxin B. These compounds typically exploit negatively charged phosphate residues on the diglucosamine backbone to bind to the cell surface and then insert into the membrane, causing disruption of the permeability barrier and lysis of the cell (Brown and Hancock, 2006). Many bacteria can overcome susceptibility to these compounds by modifying the 1 and 4’ phosphates with L-Ara4N or phosphorylethanolamine (PEtN) by the activity of ArnT and EptA respectively, following transfer of the nascent lipid A-core to the periplasm (Raetz et al., 2007). The enzymes involved in these processes in E. coli are activated under specific growth conditions (e.g. growth at low pH or in the presence of cationic peptides) and are under the regulation of the PmrAB and PhoPQ two-component systems (Guo et al., 1997 Gunn, 2008).

An additional modification involves phosphorylation of the 1-phosphate on the lipid A backbone to make pyrophosphate and is mediated by LpxT using undecaprenyl pyrophosphate as a donor (Touze et al., 2008). This activity helps recycle the essential lipid carrier for peptidoglycan, O antigen, group 1 K antigens, etc., but other cellular proteins can also fulfill this need (El Ghachi et al., 2005). The activity of LpxT is inhibited in PmrA-activated cells (Herrera et al., 2010).


Engineered K1F bacteriophages kill intracellular Escherichia coli K1 in human epithelial cells

Bacterial infections can be treated with bacteriophages that show great specificity towards their bacterial host and can be genetically modified for different applications. However, whether and how bacteriophages can kill intracellular bacteria in human cells remains elusive. Here, using CRISPR/Cas selection, we have engineered a fluorescent bacteriophage specific for E. coli K1, a nosocomial pathogen responsible for urinary tract infections, neonatal meningitis and sepsis. By confocal and live microscopy, we show that engineered bacteriophages K1F-GFP and E. coli EV36-RFP bacteria displaying the K1 capsule, enter human cells via phagocytosis. Importantly, we show that bacteriophage K1F-GFP efficiently kills intracellular E. coli EV36-RFP in T24 human urinary bladder epithelial cells. Finally, we provide evidence that bacteria and bacteriophages are degraded by LC3-associated phagocytosis and xenophagy.

Conflict of interest statement

The authors declare no competing interests.

Figures

Construction of fluorescent phages K1F.…

Construction of fluorescent phages K1F. ( A ) The engineering of three different…

Image analysis of E. coli…

Image analysis of E. coli EV36-RFP bacteria and phage K1F-GFP constructs inside epithelial…

Phage K1F targets extracellular and…

Phage K1F targets extracellular and intracellular bacteria in epithelial human cells. ( A,B…

Degradation of E. coli EV36…

Degradation of E. coli EV36 and phage K1F via LC3-assisted phagocytosis. ( A…

Phage K1F in the absence of E. coli EV36 cannot activate autophagy. (…


Biology 171


Close your eyes and picture a brick wall. What is the wall’s basic building block? It is a single brick. Like a brick wall, cells are the building blocks that make up your body.

Your body has many kinds of cells, each specialized for a specific purpose. Just as we use a variety of materials to build a home, the human body is constructed from many cell types. For example, epithelial cells protect the body’s surface and cover the organs and body cavities within. Bone cells help to support and protect the body. Immune system cells fight invading bacteria. Additionally, blood and blood cells carry nutrients and oxygen throughout the body while removing carbon dioxide. Each of these cell types plays a vital role during the body’s growth, development, and day-to-day maintenance. In spite of their enormous variety, however, cells from all organisms—even ones as diverse as bacteria, onion, and human—share certain fundamental characteristics.

Learning Objectives

By the end of this section, you will be able to do the following:

  • Name examples of prokaryotic and eukaryotic organisms
  • Compare and contrast prokaryotic and eukaryotic cells
  • Describe the relative sizes of different cells
  • Explain why cells must be small

Cells fall into one of two broad categories: prokaryotic and eukaryotic. We classify only the predominantly single-celled organisms Bacteria and Archaea as prokaryotes (pro- = “before” -kary- = “nucleus”). Animal cells, plants, fungi, and protists are all eukaryotes (eu- = “true”).

Components of Prokaryotic Cells

All cells share four common components: 1) a plasma membrane, an outer covering that separates the cell’s interior from its surrounding environment 2) cytoplasm, consisting of a jelly-like cytosol within the cell in which there are other cellular components 3) DNA, the cell’s genetic material and 4) ribosomes, which synthesize proteins. However, prokaryotes differ from eukaryotic cells in several ways.

A prokaryote is a simple, mostly single-celled (unicellular) organism that lacks a nucleus, or any other membrane-bound organelle. We will shortly come to see that this is significantly different in eukaryotes. Prokaryotic DNA is in the cell’s central part: the nucleoid ((Figure)).


Most prokaryotes have a peptidoglycan cell wall and many have a polysaccharide capsule ((Figure)). The cell wall acts as an extra layer of protection, helps the cell maintain its shape, and prevents dehydration. The capsule enables the cell to attach to surfaces in its environment. Some prokaryotes have flagella, pili, or fimbriae. Flagella are used for locomotion. Pili exchange genetic material during conjugation, the process by which one bacterium transfers genetic material to another through direct contact. Bacteria use fimbriae to attach to a host cell.

Microbiologist The most effective action anyone can take to prevent the spread of contagious illnesses is to wash his or her hands. Why? Because microbes (organisms so tiny that they can only be seen with microscopes) are ubiquitous. They live on doorknobs, money, your hands, and many other surfaces. If someone sneezes into his hand and touches a doorknob, and afterwards you touch that same doorknob, the microbes from the sneezer’s mucus are now on your hands. If you touch your hands to your mouth, nose, or eyes, those microbes can enter your body and could make you sick.

However, not all microbes (also called microorganisms) cause disease most are actually beneficial. You have microbes in your gut that make vitamin K. Other microorganisms are used to ferment beer and wine.

Microbiologists are scientists who study microbes. Microbiologists can pursue a number of careers. Not only do they work in the food industry, they are also employed in the veterinary and medical fields. They can work in the pharmaceutical sector, serving key roles in research and development by identifying new antibiotic sources that can treat bacterial infections.

Environmental microbiologists may look for new ways to use specially selected or genetically engineered microbes to remove pollutants from soil or groundwater, as well as hazardous elements from contaminated sites. We call using these microbes bioremediation technologies. Microbiologists can also work in the bioinformatics field, providing specialized knowledge and insight for designing, developing, and specificity of computer models of, for example, bacterial epidemics.

Cell Size

At 0.1 to 5.0 μm in diameter, prokaryotic cells are significantly smaller than eukaryotic cells, which have diameters ranging from 10 to 100 μm ((Figure)). The prokaryotes’ small size allows ions and organic molecules that enter them to quickly diffuse to other parts of the cell. Similarly, any wastes produced within a prokaryotic cell can quickly diffuse. This is not the case in eukaryotic cells, which have developed different structural adaptations to enhance intracellular transport.


Small size, in general, is necessary for all cells, whether prokaryotic or eukaryotic. Let’s examine why that is so. First, we’ll consider the area and volume of a typical cell. Not all cells are spherical in shape, but most tend to approximate a sphere. You may remember from your high school geometry course that the formula for the surface area of a sphere is 4πr 2 , while the formula for its volume is 4πr 3 /3. Thus, as the radius of a cell increases, its surface area increases as the square of its radius, but its volume increases as the cube of its radius (much more rapidly). Therefore, as a cell increases in size, its surface area-to-volume ratio decreases. This same principle would apply if the cell had a cube shape ((Figure)). If the cell grows too large, the plasma membrane will not have sufficient surface area to support the rate of diffusion required for the increased volume. In other words, as a cell grows, it becomes less efficient. One way to become more efficient is to divide. Another way is to develop organelles that perform specific tasks. These adaptations lead to developing more sophisticated cells, which we call eukaryotic cells.


Prokaryotic cells are much smaller than eukaryotic cells. What advantages might small cell size confer on a cell? What advantages might large cell size have?

Section Summary

Prokaryotes are single-celled organisms of the domains Bacteria and Archaea. All prokaryotes have plasma membranes, cytoplasm, ribosomes, and DNA that is not membrane-bound. Most have peptidoglycan cell walls and many have polysaccharide capsules. Prokaryotic cells range in diameter from 0.1 to 5.0 μm.

As a cell increases in size, its surface area-to-volume ratio decreases. If the cell grows too large, the plasma membrane will not have sufficient surface area to support the rate of diffusion required for the increased volume.

Art Connections

(Figure) Prokaryotic cells are much smaller than eukaryotic cells. What advantages might small cell size confer on a cell? What advantages might large cell size have?

(Figure) Substances can diffuse more quickly through small cells. Small cells have no need for organelles and therefore do not need to expend energy getting substances across organelle membranes. Large cells have organelles that can separate cellular processes, enabling them to build molecules that are more complex.

Free Response

Antibiotics are medicines that are used to fight bacterial infections. These medicines kill prokaryotic cells without harming human cells. What part or parts of the bacterial cell do you think antibiotics target? Why?

The cell wall would be targeted by antibiotics as well as the bacteria’s ability to replicate. This would inhibit the bacteria’s ability to reproduce, and it would compromise its defense mechanisms.

Explain why not all microbes are harmful.

Some microbes are beneficial. For instance, E. coli bacteria populate the human gut and help break down fiber in the diet. Some foods such as yogurt are formed by bacteria.

Glossary


Pathology

Chances are, if the spinach contained E. coli O157:H7 , he would become very ill and be in need of medical attention. Poor Popeye, don't worry Olive Oyl will take care of you!

Unfortunately , there is more than one pathogenic strain of E. coli. As stated in Interesting Facts , there are over 700 serotypes of E. coli. These are based on three different antigens: the O antigen which is derived from the cell wall, the H antigen which is derived from flagella that are used for motility, and the K antigen which is derived from a polysaccharide capsule that is secreted. Most of these specific strains can be further divided into three more categories based on how they infect the human body: Urinary Tract Infections (UTI), Neonatal Meningitis , and Intestinal Diseases ( gastroenteritis ).

1. Urinary Tract Infections

90% of UTI's are caused by Uropathogenic E. coli ( UPEC ). This is caused by E. coli colonizing in the feces and/or the perineal region, and then somehow ascending to the urethra (the urinary tract) and finally the bladder causing inflammation. The bacteria are able to adhere and aggregate to the urethra and the bladder by use of fimbriae. The movement of E. coli can occur during sexual intercourse, wiping back to front after using the restroom, among other ways, and are more common in females. Symptoms include painful urination, frequent urination, and cloudy urine. They are easily treated with antibiotics.

Neonatal Meningitis (inflammation of the meninges ), which affects 1/2000 infants, is caused by E. coli strains invading the either the nasopharynx or the GI tract, absorbed into the bloodstream, and then the blood carries the bacteria to the meninges. This is treated by antibiotic therapy, most often with ampicillin. This can be fatal if untreated.

E. coli is best known for its ability to infect the intestinal system, causing diarrhea , and these can be further categorized into five subgroups based on their pathogenicity. These diseases are treated with antibiotics.


Lab 6: Gram Stain and Capsule Stain

The Gram stain is the most widely used staining procedure in bacteriology. It is called a differential stain since it differentiates between Gram-positive and Gram-negative bacteria. Bacteria that stain purple with the Gram staining procedure are termed Gram-positive those that stain pink are said to be Gram-negative. The terms positive and negative have nothing to do with electrical charge, but simply designate two distinct morphological groups of bacteria.

Gram-positive and Gram-negative bacteria stain differently because of fundamental differences in the structure of their cell walls. The bacterial cell wall serves to give the organism its size and shape as well as to prevent osmotic lysis. The material in the bacterial cell wall which confers rigidity is peptidoglycan.

In electron micrographs, the Gram-positive cell wall appears as a broad, dense wall 20-80 nm thick and consisting of numerous interconnecting layers of peptidoglycan (Figures 1A and 1B). Chemically, 60 to 90% of the Gram-positive cell wall is peptidoglycan. Interwoven in the cell wall of Gram-positive are teichoic acids. Teichoic acids, which extend through and beyond the rest of the cell wall, are composed of polymers of glycerol, phosphates, and the sugar alcohol ribitol. Some have a lipid attached (lipoteichoic acid). The outer surface of the peptidoglycan is studded with proteins that differ with the strain and species of the bacterium.

The Gram-negative cell wall, on the other hand, contains only 2-3 layers of peptidoglycan and is surrounded by an outer membrane composed of phospholipids, lipopolysaccharide, lipoprotein, and proteins (Figures 2A and 2B). Only 10% - 20% of the Gram-negative cell wall is peptidoglycan. The phospholipids are located mainly in the inner layer of the outer membrane, as are the lipoproteins that connect the outer membrane to the peptidoglycan. The lipopolysaccharides, located in the outer layer of the outer membrane, consist of a lipid portion called lipid A embedded in the membrane and a polysaccharide portion extending outward from the bacterial surface. The outer membrane also contains a number of proteins that differ with the strain and species of the bacterium.

For further information on the Gram-negative and Gram-positive cell wall, see the following Learning Objects in your Lecture Guide:

The Gram staining procedure involves four basic steps:

1. The bacteria are first stained with the basic dye crystal violet. Both Gram-positive and Gram-negative bacteria become directly stained and appear purple after this step.

2. The bacteria are then treated with Gram's iodine solution. This allows the stain to be retained better by forming an insoluble crystal violet-iodine complex. Both Gram-positive and Gram-negative bacteria remain purple after this step.

3. Gram's decolorizer, a mixture of ethyl alcohol and acetone, is then added. This is the differential step. Gram-positive bacteria retain the crystal violet-iodine complex while Gram-negative are decolorized.

4. Finally, the counterstain safranin (also a basic dye) is applied. Since the Gram-positive bacteria are already stained purple, they are not affected by the counterstain. Gram-negative bacteria, which are now colorless, become directly stained by th e safranin. Thus, Gram-positive appear purple, and Gram-negative appear pink.

With the current theory behind Gram staining, it is thought that in Gram-positive bacteria, the crystal violet and iodine combine to form a larger molecule that precipitates out within the cell. The alcohol/acetone mixture then causes dehydration of the multilayered peptidoglycan, thus decreasing the space between the molecules and causing the cell wall to trap the crystal violet-iodine complex within the cell. In the case of Gram-negative bacteria, the alcohol/acetone mixture, being a lipid solvent, dissolves the outer membrane of the cell wall and may also damage the cytoplasmic membrane to which the peptidoglycan is attached. The few layers of peptidoglycan are unable to retain the crystal violet-iodine complex and the cell is decolorized.

It is important to note that Gram-positivity (the ability to retain the purple crystal violet-iodine complex) is not an all-or-nothing phenomenon but a matter of degree. There are several factors that could result in a Gram-positive organism staining Gram-negatively:

1. The method and techniques used. Overheating during heat fixation, over decolorization with alcohol, and even too much washing with water between steps may result in Gram-positive bacteria losing the crystal violet-iodine complex.

2. The age of the culture. Cultures more than 24 hours old may lose their ability to retain the crystal violet-iodine complex.

3. The organism itself. Some Gram-positive bacteria are more able to retain the crystal violet-iodine complex than others.

Therefore, one must use very precise techniques in Gram staining and interpret the results with discretion.

Trypticase Soy agar plate cultures of Escherichia coli (a small, Gram-negative bacillus) and Staphylococcus epidermidis (a Gram-positive coccus with a staphylococcus arrangement).

PROCEDURE (to be done individually)

1. Escherichia coli

a. Heat-fix a smear of Escherichia coli as follows:

1. Using the dropper bottle of deionized water found in your staining rack, place 1/2 of a normal sized drop of water on a clean slide by touching the dropper to the slide (Figure 1). Altenately, use your sterilized inoculating loop to place a drop of deionized water on the slide.

2. Using your sterile inoculating loop, aseptically remove a small amount of the culture from the agar surface and gently touch it 2 - 3 times to the drop of water until the water becomes visibly cloudy (Figure 2). A good smear with the correct amount of bacteria is essential to Gram staining.

  • Too many bacteria on the slide could result in under-decolorization too few could lead to over-decolorization.

3. Incinerate the remaining bacteria on the inoculating loop. If too much culture is added to the water, you will not see stained individual bacteria and you may not have a reliable Gram stain.

4 . After the inoculating loop cools, spread the suspension over approximately half of the slide to form a thin film. A correctly prepared smear with the right amount of bacteria should look similar to Fig. 3.

5 . Allow this thin suspension to completely air dry (Figure 4). The smear must be completely dry before the slide is heat fixed!

6 . To heat-fix the bacteria to the slide,pick up the air-dried slide with coverslip forceps and hold the bottom of the slide opposite the smear near the opening of the microincinerator for 10 seconds (Figure 5) as demonstrated by your instructor. If the slide is not heated enough, all of the bacteria will wash off. If it is overheated, the bacteria structural integrity can be damaged.

b . Stain with Hucker's crystal violet for one minute (Figure 6). Gently wash with water (Figure 7). Shake off the excess water but do not blot dry between steps.

c . Stain with Gram's iodine solution for one minute (Figure 8) and gently wash with water.

d . Decolorize by picking up the slide and letting the Gram's decolorizer run down the slide until the purple just stops flowing at the bottom of the slide (Figure 9).

  • Make sure the entire smear is evenly decolorized and that you are not under-decolorizing or over-decolorizing.
  • Wash immediately with water.

e . Stain with safranin for one minute (Figure 10). When you wash off the excess safranin, be very careful to wash gently and briefly as it is possible to wash out some of the sarfanin in the bacterium.

f . Blot dry (Figure 11) and observe using oil immersion microscopy.

2 . Staphylococcus epidermidis

a. Heat-fix a smear of Staphylococcus epidermidis as follows:

1. Using the dropper bottle of deionized water found in your staining rack, place 1/2 of a normal sized drop of water on a clean slide by touching the dropper to the slide (Figure 1). Altenately, use your sterilized inoculating loop to place a drop of deionized water on the slide.

2. Using your sterile inoculating loop, aseptically remove a small amount of the culture from the agar surface and gently touch it 2 - 3 times to the drop of water until the water becomes visibly cloudy (Figure 2).

  • Too many bacteria on the slide could result in under-decolorization too few could lead to over-decolorization.

3. Incinerate the remaining bacteria on the inoculating loop. If too much culture is added to the water, you will not see stained individual bacteria and you may not have a reliable Gram stain.

4 . After the inoculating loop cools, spread the suspension over approximately half of the slide to form a thin film (Figure 3).

5 . Allow this thin suspension to completely air dry (Figure 4). The smear must be completely dry before the slide is heat fixed!

6 . To heat-fix the bacteria to the slide,pick up the air-dried slide with coverslip forceps and hold the bottom of the slide opposite the smear near the opening of the microincinerator for 10 seconds (Figure 5) as demonstrated by your instructor. If the slide is not heated enough, all of the bacteria will wash off. If it is overheated, the bacteria structural integrity can be damaged.

b . Stain with Hucker's crystal violet for one minute (Figure 6). Gently wash with water (Figure 7). Shake off the excess water but do not blot dry between steps.

c . Stain with Gram's iodine solution for one minute (Figure 8) and gently wash with water.

d . Decolorize by picking up the slide and letting the Gram's decolorizer run down the slide until the purple just stops flowing at the bottom of the slide (Figure 9).

  • Make sure the entire smear is evenly decolorized and that you are not under-decolorizing or over-decolorizing.
  • Wash immediately with water.

e . Stain with safranin for one minute (Figure 10). When you wash off the excess safranin, be very careful to wash gently and briefly as it is possible to wash out some of the sarfanin in the bacterium.

f . Blot dry and observe using oil immersion microscopy.

3. Make sure you carefully pour the used dye in your staining tray into the waste dye collection container, not down the sink.

B. THE CAPSULE STAIN

Many bacteria secrete a slimy, viscous covering called a capsule or glycocalyx . This is usually composed of polysaccharide, polypeptide, or both.

The ability to produce a capsule is an inherited property of the organism, but the capsule is not an absolutely essential cellular component. Capsules are often produced only under specific growth conditions. Even though not essential for life, capsules p robably help bacteria to survive in nature. Capsules help many pathogenic and normal flora bacteria to initially resist phagocytosis by the host's phagocytic cells. In soil and water, capsules help prevent bacteria from being engulfed by protozoans. Capsules also help many bacteria to adhere to surfaces and thus resist flushing. It also enables many bacteria to form biofilms. A biofilm consists layers of bacterial populations adhering to host cells and embedded in a common capsular mass.

For further information on the bacterial capsules, see the following Learning Objects in your Lecture Guide:

Skim Milk broth culture of Enterobacter aerogenes. The skim milk supplies essential nutrients for capsule production and also provides a slightly stainable background.

PROCEDURE (to be done individually)

1. Stir up the Skim Milk broth culture with your loop and place 2-3 loops of Enterobacter aerogenes on a microscope slide.

2. Using your inoculating loop, spread the sample out to cover about one inch of the slide.

3. Let it completely air dry. Do not heat fix. Capsules stick well to glass, and heat may destroy the capsule.

4. Stain with crystal violet for one minute.

5. Wash off the excess dye with 20% copper sulfate solution.

6. Shake off the excess copper sulfate solution and immediately blot dry.

7. Observe using oil immersion microscopy. The organism and the milk dried on the slide will pick up the purple dye while the capsule will remain colorless.

8 . Make sure you carefully pour the used dye in your staining tray into the waste dye collection container, not down the sink.

9. Observe the demonstration capsule stain of Streptococcus lactis , an encapsulated bacterium that is normal flora in milk.


Contents

Type and morphology Edit

E. coli is a Gram-negative, facultative anaerobe (that makes ATP by aerobic respiration if oxygen is present, but is capable of switching to fermentation or anaerobic respiration if oxygen is absent) and nonsporulating bacterium. [17] Cells are typically rod-shaped, and are about 2.0 μm long and 0.25–1.0 μm in diameter, with a cell volume of 0.6–0.7 μm 3 . [18] [19] [20]

E. coli stains Gram-negative because its cell wall is composed of a thin peptidoglycan layer and an outer membrane. During the staining process, E. coli picks up the color of the counterstain safranin and stains pink. The outer membrane surrounding the cell wall provides a barrier to certain antibiotics such that E. coli is not damaged by penicillin. [15]

Strains that possess flagella are motile. The flagella have a peritrichous arrangement. [21] It also attaches and effaces to the microvilli of the intestines via an adhesion molecule known as intimin. [22]

Metabolism Edit

E. coli can live on a wide variety of substrates and uses mixed acid fermentation in anaerobic conditions, producing lactate, succinate, ethanol, acetate, and carbon dioxide. Since many pathways in mixed-acid fermentation produce hydrogen gas, these pathways require the levels of hydrogen to be low, as is the case when E. coli lives together with hydrogen-consuming organisms, such as methanogens or sulphate-reducing bacteria. [23]

In addition, E. coli's metabolism can be rewired to solely use CO2 as the source of carbon for biomass production. In other words, this obligate heterotroph's metabolism can be altered to display autotrophic capabilities by heterologously expressing carbon fixation genes as well as formate dehydrogenase and conducting laboratory evolution experiments. This may be done by using formate to reduce electron carriers and supply the ATP required in anabolic pathways inside of these synthetic autotrophs. [24]

E. coli have three native glycolytic pathways: EMPP, EDP, and OPPP. The EMPP employs ten enzymatic steps to yield two pyruvates, two ATP, and two NADH per glucose molecule while OPPP serves as an oxidation route for NADPH synthesis. Although the EDP is the more thermodynamically favorable of the three pathways, E. coli do not use the EDP for glucose metabolism, relying mainly on the EMPP and the OPPP. The EDP mainly remains inactive except for during growth with gluconate. [25]

Catabolite Repression Edit

When growing in the presence of a mixture of sugars, bacteria will often consume the sugars sequentially through a process known as catabolite repression. By repressing the expression of the genes involved in metabolizing the less preferred sugars, cells will usually first consume the sugar yielding the highest growth rate, followed by the sugar yielding the next highest growth rate, and so on. In doing so the cells ensure that their limited metabolic resources are being used to maximize the rate of growth. The well-used example of this with E. coli involves the growth of the bacterium on glucose and lactose, where E. coli will consume glucose before lactose. Catabolite repression has also been observed in E.coli in the presence of other non-glucose sugars, such as arabinose and xylose, sorbitol, rhamnose, and ribose. In E. coli, glucose catabolite repression is regulated by the phosphotransferase system, a multi-protein phosphorylation cascade that couples glucose uptake and metabolism. [26]

Culture growth Edit

Optimum growth of E. coli occurs at 37 °C (98.6 °F), but some laboratory strains can multiply at temperatures up to 49 °C (120 °F). [27] E. coli grows in a variety of defined laboratory media, such as lysogeny broth, or any medium that contains glucose, ammonium phosphate monobasic, sodium chloride, magnesium sulfate, potassium phosphate dibasic, and water. Growth can be driven by aerobic or anaerobic respiration, using a large variety of redox pairs, including the oxidation of pyruvic acid, formic acid, hydrogen, and amino acids, and the reduction of substrates such as oxygen, nitrate, fumarate, dimethyl sulfoxide, and trimethylamine N-oxide. [28] E. coli is classified as a facultative anaerobe. It uses oxygen when it is present and available. It can, however, continue to grow in the absence of oxygen using fermentation or anaerobic respiration. The ability to continue growing in the absence of oxygen is an advantage to bacteria because their survival is increased in environments where water predominates. [15]

Cell cycle Edit

The bacterial cell cycle is divided into three stages. The B period occurs between the completion of cell division and the beginning of DNA replication. The C period encompasses the time it takes to replicate the chromosomal DNA. The D period refers to the stage between the conclusion of DNA replication and the end of cell division. [29] The doubling rate of E. coli is higher when more nutrients are available. However, the length of the C and D periods do not change, even when the doubling time becomes less than the sum of the C and D periods. At the fastest growth rates, replication begins before the previous round of replication has completed, resulting in multiple replication forks along the DNA and overlapping cell cycles. [30]

The number of replication forks in fast growing E. coli typically follows 2n (n = 1, 2 or 3). This only happens if replication is initiated simultaneously from all origins of replications, and is referred to as synchronous replication. However, not all cells in a culture replicate synchronously. In this case cells do not have multiples of two replication forks. Replication initiation is then referred to being asynchronous. [31] However, asynchrony can be caused by mutations to for instance DnaA [31] or DnaA initiator-associating protein DiaA. [32]

Genetic adaptation Edit

E. coli and related bacteria possess the ability to transfer DNA via bacterial conjugation or transduction, which allows genetic material to spread horizontally through an existing population. The process of transduction, which uses the bacterial virus called a bacteriophage, [33] is where the spread of the gene encoding for the Shiga toxin from the Shigella bacteria to E. coli helped produce E. coli O157:H7, the Shiga toxin-producing strain of E. coli.

E. coli encompasses an enormous population of bacteria that exhibit a very high degree of both genetic and phenotypic diversity. Genome sequencing of many isolates of E. coli and related bacteria shows that a taxonomic reclassification would be desirable. However, this has not been done, largely due to its medical importance, [34] and E. coli remains one of the most diverse bacterial species: only 20% of the genes in a typical E. coli genome is shared among all strains. [35]

In fact, from the more constructive point of view, the members of genus Shigella (S. dysenteriae, S. flexneri, S. boydii, and S. sonnei) should be classified as E. coli strains, a phenomenon termed taxa in disguise. [36] Similarly, other strains of E. coli (e.g. the K-12 strain commonly used in recombinant DNA work) are sufficiently different that they would merit reclassification.

A strain is a subgroup within the species that has unique characteristics that distinguish it from other strains. These differences are often detectable only at the molecular level however, they may result in changes to the physiology or lifecycle of the bacterium. For example, a strain may gain pathogenic capacity, the ability to use a unique carbon source, the ability to take upon a particular ecological niche, or the ability to resist antimicrobial agents. Different strains of E. coli are often host-specific, making it possible to determine the source of fecal contamination in environmental samples. [12] [13] For example, knowing which E. coli strains are present in a water sample allows researchers to make assumptions about whether the contamination originated from a human, another mammal, or a bird.

Serotypes Edit

A common subdivision system of E. coli, but not based on evolutionary relatedness, is by serotype, which is based on major surface antigens (O antigen: part of lipopolysaccharide layer H: flagellin K antigen: capsule), e.g. O157:H7). [37] It is, however, common to cite only the serogroup, i.e. the O-antigen. At present, about 190 serogroups are known. [38] The common laboratory strain has a mutation that prevents the formation of an O-antigen and is thus not typeable.

Genome plasticity and evolution Edit

Like all lifeforms, new strains of E. coli evolve through the natural biological processes of mutation, gene duplication, and horizontal gene transfer in particular, 18% of the genome of the laboratory strain MG1655 was horizontally acquired since the divergence from Salmonella. [39] E. coli K-12 and E. coli B strains are the most frequently used varieties for laboratory purposes. Some strains develop traits that can be harmful to a host animal. These virulent strains typically cause a bout of diarrhea that is often self-limiting in healthy adults but is frequently lethal to children in the developing world. [40] More virulent strains, such as O157:H7, cause serious illness or death in the elderly, the very young, or the immunocompromised. [40] [41]

The genera Escherichia and Salmonella diverged around 102 million years ago (credibility interval: 57–176 mya), which coincides with the divergence of their hosts: the former being found in mammals and the latter in birds and reptiles. [42] This was followed by a split of an Escherichia ancestor into five species (E. albertii, E. coli, E. fergusonii, E. hermannii, and E. vulneris). The last E. coli ancestor split between 20 and 30 million years ago. [43]

The long-term evolution experiments using E. coli, begun by Richard Lenski in 1988, have allowed direct observation of genome evolution over more than 65,000 generations in the laboratory. [44] For instance, E. coli typically do not have the ability to grow aerobically with citrate as a carbon source, which is used as a diagnostic criterion with which to differentiate E. coli from other, closely, related bacteria such as Salmonella. In this experiment, one population of E. coli unexpectedly evolved the ability to aerobically metabolize citrate, a major evolutionary shift with some hallmarks of microbial speciation.

In the microbial world, a relationship of predation can be established similar to that observed in the animal world. Considered, it has been seen that E. coli is the prey of multiple generalist predators, such as Myxococcus xanthus. In this predator-prey relationship, a parallel evolution of both species is observed through genomic and phenotypic modifications, in the case of E. coli the modifications are modified in two aspects involved in their virulence such as mucoid production (excessive production of exoplasmic acid alginate ) and the suppression of the OmpT gene, producing in future generations a better adaptation of one of the species that is counteracted by the evolution of the other, following a co-evolutionary model demonstrated by the Red Queen hypothesis. [45]

Neotype strain Edit

E. coli is the type species of the genus (Escherichia) and in turn Escherichia is the type genus of the family Enterobacteriaceae, where the family name does not stem from the genus Enterobacter + "i" (sic.) + "aceae", but from "enterobacterium" + "aceae" (enterobacterium being not a genus, but an alternative trivial name to enteric bacterium). [46] [47]

The original strain described by Escherich is believed to be lost, consequently a new type strain (neotype) was chosen as a representative: the neotype strain is U5/41 T , [48] also known under the deposit names DSM 30083, [49] ATCC 11775, [50] and NCTC 9001, [51] which is pathogenic to chickens and has an O1:K1:H7 serotype. [52] However, in most studies, either O157:H7, K-12 MG1655, or K-12 W3110 were used as a representative E. coli. The genome of the type strain has only lately been sequenced. [48]

Phylogeny of E. coli strains Edit

Many strains belonging to this species have been isolated and characterised. In addition to serotype (vide supra), they can be classified according to their phylogeny, i.e. the inferred evolutionary history, as shown below where the species is divided into six groups. [53] [54] Particularly the use of whole genome sequences yields highly supported phylogenies. Based on such data, five subspecies of E. coli were distinguished. [48]

The link between phylogenetic distance ("relatedness") and pathology is small, [48] e.g. the O157:H7 serotype strains, which form a clade ("an exclusive group")—group E below—are all enterohaemorragic strains (EHEC), but not all EHEC strains are closely related. In fact, four different species of Shigella are nested among E. coli strains (vide supra), while E. albertii and E. fergusonii are outside this group. Indeed, all Shigella species were placed within a single subspecies of E. coli in a phylogenomic study that included the type strain, [48] and for this reason an according reclassification is difficult. All commonly used research strains of E. coli belong to group A and are derived mainly from Clifton's K-12 strain (λ + F + O16) and to a lesser degree from d'Herelle's Bacillus coli strain (B strain)(O7).

E. coli S88 (O45:K1. Extracellular pathogenic)

E. coli UMN026 (O17:K52:H18. Extracellular pathogenic)

E. coli (O19:H34. Extracellular pathogenic)

E. coli (O7:K1. Extracellular pathogenic)

E. coli GOS1 (O104:H4 EAHEC) German 2011 outbreak

E. coli ATCC8739 (O146. Crook's E.coli used in phage work in the 1950s)

E. coli K-12 W3110 (O16. λ − F − "wild type" molecular biology strain)

E. coli K-12 DH10b (O16. high electrocompetency molecular biology strain)

E. coli K-12 DH1 (O16. high chemical competency molecular biology strain)

E. coli K-12 MG1655 (O16. λ − F − "wild type" molecular biology strain)

E. coli BW2952 (O16. competent molecular biology strain)

E. coli B REL606 (O7. high competency molecular biology strain)

E. coli BL21-DE3 (O7. expression molecular biology strain with T7 polymerase for pET system)

The first complete DNA sequence of an E. coli genome (laboratory strain K-12 derivative MG1655) was published in 1997. It is a circular DNA molecule 4.6 million base pairs in length, containing 4288 annotated protein-coding genes (organized into 2584 operons), seven ribosomal RNA (rRNA) operons, and 86 transfer RNA (tRNA) genes. Despite having been the subject of intensive genetic analysis for about 40 years, many of these genes were previously unknown. The coding density was found to be very high, with a mean distance between genes of only 118 base pairs. The genome was observed to contain a significant number of transposable genetic elements, repeat elements, cryptic prophages, and bacteriophage remnants. [55]

More than three hundred complete genomic sequences of Escherichia and Shigella species are known. The genome sequence of the type strain of E. coli was added to this collection before 2014. [48] Comparison of these sequences shows a remarkable amount of diversity only about 20% of each genome represents sequences present in every one of the isolates, while around 80% of each genome can vary among isolates. [35] Each individual genome contains between 4,000 and 5,500 genes, but the total number of different genes among all of the sequenced E. coli strains (the pangenome) exceeds 16,000. This very large variety of component genes has been interpreted to mean that two-thirds of the E. coli pangenome originated in other species and arrived through the process of horizontal gene transfer. [56]

Genes in E. coli are usually named by 4-letter acronyms that derive from their function (when known) and italicized. For instance, recA is named after its role in homologous recombination plus the letter A. Functionally related genes are named recB, recC, recD etc. The proteins are named by uppercase acronyms, e.g. RecA, RecB, etc. When the genome of E. coli was sequenced, all genes were numbered (more or less) in their order on the genome and abbreviated by b numbers, such as b2819 (= recD). The "b" names were created after Fred Blattner, who led the genome sequence effort. [55] Another numbering system was introduced with the sequence of another E. coli strain, W3110, which was sequenced in Japan and hence uses numbers starting by JW. (Japanese W3110), e.g. JW2787 (= recD). [57] Hence, recD = b2819 = JW2787. Note, however, that most databases have their own numbering system, e.g. the EcoGene database [58] uses EG10826 for recD. Finally, ECK numbers are specifically used for alleles in the MG1655 strain of E. coli K-12. [58] Complete lists of genes and their synonyms can be obtained from databases such as EcoGene or Uniprot.

Proteome Edit

Several studies have investigated the proteome of E. coli. By 2006, 1,627 (38%) of the 4,237 open reading frames (ORFs) had been identified experimentally. [59] The 4,639,221–base pair sequence of Escherichia coli K-12 is presented. Of 4288 protein-coding genes annotated, 38 percent have no attributed function. Comparison with five other sequenced microbes reveals ubiquitous as well as narrowly distributed gene families many families of similar genes within E. coli are also evident. The largest family of paralogous proteins contains 80 ABC transporters. The genome as a whole is strikingly organized with respect to the local direction of replication guanines, oligonucleotides possibly related to replication and recombination, and most genes are so oriented. The genome also contains insertion sequence (IS) elements, phage remnants, and many other patches of unusual composition indicating genome plasticity through horizontal transfer. [55]

Interactome Edit

The interactome of E. coli has been studied by affinity purification and mass spectrometry (AP/MS) and by analyzing the binary interactions among its proteins.

Protein complexes. A 2006 study purified 4,339 proteins from cultures of strain K-12 and found interacting partners for 2,667 proteins, many of which had unknown functions at the time. [60] A 2009 study found 5,993 interactions between proteins of the same E. coli strain, though these data showed little overlap with those of the 2006 publication. [61]

Binary interactions. Rajagopala et al. (2014) have carried out systematic yeast two-hybrid screens with most E. coli proteins, and found a total of 2,234 protein-protein interactions. [62] This study also integrated genetic interactions and protein structures and mapped 458 interactions within 227 protein complexes.

E. coli belongs to a group of bacteria informally known as coliforms that are found in the gastrointestinal tract of warm-blooded animals. [63] E. coli normally colonizes an infant's gastrointestinal tract within 40 hours of birth, arriving with food or water or from the individuals handling the child. In the bowel, E. coli adheres to the mucus of the large intestine. It is the primary facultative anaerobe of the human gastrointestinal tract. [64] (Facultative anaerobes are organisms that can grow in either the presence or absence of oxygen.) As long as these bacteria do not acquire genetic elements encoding for virulence factors, they remain benign commensals. [65]

Therapeutic use Edit

Due to the low cost and speed with which it can be grown and modified in laboratory settings, E. coli is a popular expression platform for the production of recombinant proteins used in therapeutics. One advantage to using E. coli over another expression platform is that E. coli naturally does not export many proteins into the periplasm, making it easier to recover a protein of interest without cross-contamination. [66] The E. coli K-12 strains and their derivatives (DH1, DH5α, MG1655, RV308 and W3110) are the strains most widely used by the biotechnology industry. [67] Nonpathogenic E. coli strain Nissle 1917 (EcN), (Mutaflor) and E. coli O83:K24:H31 (Colinfant) [68] [69] ) are used as probiotic agents in medicine, mainly for the treatment of various gastrointestinal diseases, [70] including inflammatory bowel disease. [71] It is thought that the EcN strain might impede the growth of opportunistic pathogens, including Salmonella and other coliform enteropathogens, through the production of microcin proteins the production of siderophores. [72]

Most E. coli strains do not cause disease, naturally living in the gut, [73] but virulent strains can cause gastroenteritis, urinary tract infections, neonatal meningitis, hemorrhagic colitis, and Crohn's disease. Common signs and symptoms include severe abdominal cramps, diarrhea, hemorrhagic colitis, vomiting, and sometimes fever. In rarer cases, virulent strains are also responsible for bowel necrosis (tissue death) and perforation without progressing to hemolytic-uremic syndrome, peritonitis, mastitis, sepsis, and Gram-negative pneumonia. Very young children are more susceptible to develop severe illness, such as hemolytic uremic syndrome however, healthy individuals of all ages are at risk to the severe consequences that may arise as a result of being infected with E. coli. [64] [74] [75] [76]

Some strains of E. coli, for example O157:H7, can produce Shiga toxin (classified as a bioterrorism agent). The Shiga toxin causes inflammatory responses in target cells of the gut, leaving behind lesions which result in the bloody diarrhea that is a symptom of a Shiga toxin-producing E. coli (STEC) infection. This toxin further causes premature destruction of the red blood cells, which then clog the body's filtering system, the kidneys, in some rare cases (usually in children and the elderly) causing hemolytic-uremic syndrome (HUS), which may lead to kidney failure and even death. Signs of hemolytic uremic syndrome include decreased frequency of urination, lethargy, and paleness of cheeks and inside the lower eyelids. In 25% of HUS patients, complications of nervous system occur, which in turn causes strokes. In addition, this strain causes the buildup of fluid (since the kidneys do not work), leading to edema around the lungs, legs, and arms. This increase in fluid buildup especially around the lungs impedes the functioning of the heart, causing an increase in blood pressure. [77] [22] [78] [79] [80] [75] [76]

Uropathogenic E. coli (UPEC) is one of the main causes of urinary tract infections. [81] It is part of the normal microbiota in the gut and can be introduced in many ways. In particular for females, the direction of wiping after defecation (wiping back to front) can lead to fecal contamination of the urogenital orifices. Anal intercourse can also introduce this bacterium into the male urethra, and in switching from anal to vaginal intercourse, the male can also introduce UPEC to the female urogenital system.

Enterotoxigenic E. coli (ETEC) is the most common cause of traveler's diarrhea, with as many as 840 million cases worldwide in developing countries each year. The bacteria, typically transmitted through contaminated food or drinking water, adheres to the intestinal lining, where it secretes either of two types of enterotoxins, leading to watery diarrhea. The rate and severity of infections are higher among children under the age of five, including as many as 380,000 deaths annually. [82]

In May 2011, one E. coli strain, O104:H4, was the subject of a bacterial outbreak that began in Germany. Certain strains of E. coli are a major cause of foodborne illness. The outbreak started when several people in Germany were infected with enterohemorrhagic E. coli (EHEC) bacteria, leading to hemolytic-uremic syndrome (HUS), a medical emergency that requires urgent treatment. The outbreak did not only concern Germany, but also 15 other countries, including regions in North America. [83] On 30 June 2011, the German Bundesinstitut für Risikobewertung (BfR) (Federal Institute for Risk Assessment, a federal institute within the German Federal Ministry of Food, Agriculture and Consumer Protection) announced that seeds of fenugreek from Egypt were likely the cause of the EHEC outbreak. [84]

Some studies have demonstrated an absence of E.coli in the gut flora of subjects with the metabolic disorder Phenylketonuria. It is hypothesized that the absence of these normal bacterium impairs the production of the key vitamins B2 (riboflavin) and K2 (menaquinone) - vitamins which are implicated in many physiological roles in humans such as cellular and bone metabolism - and so contributes to the disorder. [85]

Incubation period Edit

The time between ingesting the STEC bacteria and feeling sick is called the "incubation period". The incubation period is usually 3–4 days after the exposure, but may be as short as 1 day or as long as 10 days. The symptoms often begin slowly with mild belly pain or non-bloody diarrhea that worsens over several days. HUS, if it occurs, develops an average 7 days after the first symptoms, when the diarrhea is improving. [86]

Diagnosis Edit

Diagnosis of infectious diarrhea and identification of antimicrobial resistance is performed using a stool culture with subsequent antibiotic sensitivity testing. It requires a minimum of 2 days and maximum of several weeks to culture gastrointestinal pathogens. The sensitivity (true positive) and specificity (true negative) rates for stool culture vary by pathogen, although a number of human pathogens can not be cultured. For culture-positive samples, antimicrobial resistance testing takes an additional 12-24 hours to perform.

Current point of care molecular diagnostic tests can identify E. coli and antimicrobial resistance in the identified strains much faster than culture and sensitivity testing. Microarray-based platforms can identify specific pathogenic strains of E. coli and E. coli-specific AMR genes in two hours or less with high sensitivity and specificity, but the size of the test panel (i.e., total pathogens and antimicrobial resistance genes) is limited. Newer metagenomics-based infectious disease diagnostic platforms are currently being developed to overcome the various limitations of culture and all currently available molecular diagnostic technologies.

Treatment Edit

The mainstay of treatment is the assessment of dehydration and replacement of fluid and electrolytes. Administration of antibiotics has been shown to shorten the course of illness and duration of excretion of enterotoxigenic E. coli (ETEC) in adults in endemic areas and in traveller's diarrhea, though the rate of resistance to commonly used antibiotics is increasing and they are generally not recommended. [87] The antibiotic used depends upon susceptibility patterns in the particular geographical region. Currently, the antibiotics of choice are fluoroquinolones or azithromycin, with an emerging role for rifaximin. Oral rifaximin, a semisynthetic rifamycin derivative, is an effective and well-tolerated antibacterial for the management of adults with non-invasive traveller's diarrhea. Rifaximin was significantly more effective than placebo and no less effective than ciprofloxacin in reducing the duration of diarrhea. While rifaximin is effective in patients with E. coli-predominant traveller's diarrhea, it appears ineffective in patients infected with inflammatory or invasive enteropathogens. [88]

Prevention Edit

ETEC is the type of E. coli that most vaccine development efforts are focused on. Antibodies against the LT and major CFs of ETEC provide protection against LT-producing, ETEC-expressing homologous CFs. Oral inactivated vaccines consisting of toxin antigen and whole cells, i.e. the licensed recombinant cholera B subunit (rCTB)-WC cholera vaccine Dukoral, have been developed. There are currently no licensed vaccines for ETEC, though several are in various stages of development. [89] In different trials, the rCTB-WC cholera vaccine provided high (85–100%) short-term protection. An oral ETEC vaccine candidate consisting of rCTB and formalin inactivated E. coli bacteria expressing major CFs has been shown in clinical trials to be safe, immunogenic, and effective against severe diarrhoea in American travelers but not against ETEC diarrhoea in young children in Egypt. A modified ETEC vaccine consisting of recombinant E. coli strains over-expressing the major CFs and a more LT-like hybrid toxoid called LCTBA, are undergoing clinical testing. [90] [91]

Other proven prevention methods for E. coli transmission include handwashing and improved sanitation and drinking water, as transmission occurs through fecal contamination of food and water supplies. Additionally, thoroughly cooking meat and avoiding consumption of raw, unpasteurized beverages, such as juices and milk are other proven methods for preventing E.coli. Lastly, avoid cross-contamination of utensils and work spaces when preparing food. [92]

Because of its long history of laboratory culture and ease of manipulation, E. coli plays an important role in modern biological engineering and industrial microbiology. [93] The work of Stanley Norman Cohen and Herbert Boyer in E. coli, using plasmids and restriction enzymes to create recombinant DNA, became a foundation of biotechnology. [94]

E. coli is a very versatile host for the production of heterologous proteins, [95] and various protein expression systems have been developed which allow the production of recombinant proteins in E. coli. Researchers can introduce genes into the microbes using plasmids which permit high level expression of protein, and such protein may be mass-produced in industrial fermentation processes. One of the first useful applications of recombinant DNA technology was the manipulation of E. coli to produce human insulin. [96]

Many proteins previously thought difficult or impossible to be expressed in E. coli in folded form have been successfully expressed in E. coli. For example, proteins with multiple disulphide bonds may be produced in the periplasmic space or in the cytoplasm of mutants rendered sufficiently oxidizing to allow disulphide-bonds to form, [97] while proteins requiring post-translational modification such as glycosylation for stability or function have been expressed using the N-linked glycosylation system of Campylobacter jejuni engineered into E. coli. [98] [99] [100]

Modified E. coli cells have been used in vaccine development, bioremediation, production of biofuels, [101] lighting, and production of immobilised enzymes. [95] [102]

Strain K-12 is a mutant form of E. coli that over-expresses the enzyme Alkaline Phosphatase (ALP). [103] The mutation arises due to a defect in the gene that constantly codes for the enzyme. A gene that is producing a product without any inhibition is said to have constitutive activity. This particular mutant form is used to isolate and purify the aforementioned enzyme. [103]

Strain OP50 of Escherichia coli is used for maintenance of Caenorhabditis elegans cultures.

Strain JM109 is a mutant form of E. coli that is recA and endA deficient. The strain can be utilized for blue/white screening when the cells carry the fertility factor episome [104] Lack of recA decreases the possibility of unwanted restriction of the DNA of interest and lack of endA inhibit plasmid DNA decomposition. Thus, JM109 is useful for cloning and expression systems.

Model organism Edit

E. coli is frequently used as a model organism in microbiology studies. Cultivated strains (e.g. E. coli K12) are well-adapted to the laboratory environment, and, unlike wild-type strains, have lost their ability to thrive in the intestine. Many laboratory strains lose their ability to form biofilms. [105] [106] These features protect wild-type strains from antibodies and other chemical attacks, but require a large expenditure of energy and material resources. E. coli is often used as a representative microorganism in the research of novel water treatment and sterilisation methods, including photocatalysis. By standard plate count methods, following sequential dilutions, and growth on agar gel plates, the concentration of viable organisms or CFUs (Colony Forming Units), in a known volume of treated water can be evaluated, allowing the comparative assessment of materials performance. [107]

In 1946, Joshua Lederberg and Edward Tatum first described the phenomenon known as bacterial conjugation using E. coli as a model bacterium, [108] and it remains the primary model to study conjugation. [109] E. coli was an integral part of the first experiments to understand phage genetics, [110] and early researchers, such as Seymour Benzer, used E. coli and phage T4 to understand the topography of gene structure. [111] Prior to Benzer's research, it was not known whether the gene was a linear structure, or if it had a branching pattern. [112]

E. coli was one of the first organisms to have its genome sequenced the complete genome of E. coli K12 was published by Science in 1997 [55]

From 2002 to 2010, a team at the Hungarian Academy of Science created a strain of Escherichia coli called MDS42, which is now sold by Scarab Genomics of Madison, WI under the name of "Clean Genome. E.coli", [113] where 15% of the genome of the parental strain (E. coli K-12 MG1655) were removed to aid in molecular biology efficiency, removing IS elements, pseudogenes and phages, resulting in better maintenance of plasmid-encoded toxic genes, which are often inactivated by transposons. [114] [115] [116] Biochemistry and replication machinery were not altered.

By evaluating the possible combination of nanotechnologies with landscape ecology, complex habitat landscapes can be generated with details at the nanoscale. [117] On such synthetic ecosystems, evolutionary experiments with E. coli have been performed to study the spatial biophysics of adaptation in an island biogeography on-chip.

Studies are also being performed attempting to program E. coli to solve complicated mathematics problems, such as the Hamiltonian path problem. [118]

In other studies, non-pathogenic E. coli has been used as a model microorganism towards understanding the effects of simulated microgravity (on Earth) on the same. [119] [120]

In 1885, the German-Austrian pediatrician Theodor Escherich discovered this organism in the feces of healthy individuals. He called it Bacterium coli commune because it is found in the colon. Early classifications of prokaryotes placed these in a handful of genera based on their shape and motility (at that time Ernst Haeckel's classification of bacteria in the kingdom Monera was in place). [91] [121] [122]

Bacterium coli was the type species of the now invalid genus Bacterium when it was revealed that the former type species ("Bacterium triloculare") was missing. [123] Following a revision of Bacterium, it was reclassified as Bacillus coli by Migula in 1895 [124] and later reclassified in the newly created genus Escherichia, named after its original discoverer. [125]

In 1996, the world's worst to date outbreak of E. coli food poisoning occurred in Wishaw, Scotland, killing 21 people. [126] This death toll was exceeded in 2011, when the 2011 Germany E. coli O104:H4 outbreak, linked to organic fenugreek sprouts, killed 53 people.


16 Prokaryotic Cells

By the end of this section, you will be able to do the following:

  • Name examples of prokaryotic and eukaryotic organisms
  • Compare and contrast prokaryotic and eukaryotic cells
  • Describe the relative sizes of different cells
  • Explain why cells must be small

Cells fall into one of two broad categories: prokaryotic and eukaryotic. We classify only the predominantly single-celled organisms Bacteria and Archaea as prokaryotes (pro- = “before” -kary- = “nucleus”). Animal cells, plants, fungi, and protists are all eukaryotes (eu- = “true”).

Components of Prokaryotic Cells

All cells share four common components: 1) a plasma membrane, an outer covering that separates the cell’s interior from its surrounding environment 2) cytoplasm, consisting of a jelly-like cytosol within the cell in which there are other cellular components 3) DNA, the cell’s genetic material and 4) ribosomes, which synthesize proteins. However, prokaryotes differ from eukaryotic cells in several ways.

A prokaryote is a simple, mostly single-celled (unicellular) organism that lacks a nucleus, or any other membrane-bound organelle. We will shortly come to see that this is significantly different in eukaryotes. Prokaryotic DNA is in the cell’s central part: the nucleoid ((Figure)).


Most prokaryotes have a peptidoglycan cell wall and many have a polysaccharide capsule ((Figure)). The cell wall acts as an extra layer of protection, helps the cell maintain its shape, and prevents dehydration. The capsule enables the cell to attach to surfaces in its environment. Some prokaryotes have flagella, pili, or fimbriae. Flagella are used for locomotion. Pili exchange genetic material during conjugation, the process by which one bacterium transfers genetic material to another through direct contact. Bacteria use fimbriae to attach to a host cell.

Microbiologist The most effective action anyone can take to prevent the spread of contagious illnesses is to wash his or her hands. Why? Because microbes (organisms so tiny that they can only be seen with microscopes) are ubiquitous. They live on doorknobs, money, your hands, and many other surfaces. If someone sneezes into his hand and touches a doorknob, and afterwards you touch that same doorknob, the microbes from the sneezer’s mucus are now on your hands. If you touch your hands to your mouth, nose, or eyes, those microbes can enter your body and could make you sick.

However, not all microbes (also called microorganisms) cause disease most are actually beneficial. You have microbes in your gut that make vitamin K. Other microorganisms are used to ferment beer and wine.

Microbiologists are scientists who study microbes. Microbiologists can pursue a number of careers. Not only do they work in the food industry, they are also employed in the veterinary and medical fields. They can work in the pharmaceutical sector, serving key roles in research and development by identifying new antibiotic sources that can treat bacterial infections.

Environmental microbiologists may look for new ways to use specially selected or genetically engineered microbes to remove pollutants from soil or groundwater, as well as hazardous elements from contaminated sites. We call using these microbes bioremediation technologies. Microbiologists can also work in the bioinformatics field, providing specialized knowledge and insight for designing, developing, and specificity of computer models of, for example, bacterial epidemics.

Cell Size

At 0.1 to 5.0 μm in diameter, prokaryotic cells are significantly smaller than eukaryotic cells, which have diameters ranging from 10 to 100 μm ((Figure)). The prokaryotes’ small size allows ions and organic molecules that enter them to quickly diffuse to other parts of the cell. Similarly, any wastes produced within a prokaryotic cell can quickly diffuse. This is not the case in eukaryotic cells, which have developed different structural adaptations to enhance intracellular transport.


Small size, in general, is necessary for all cells, whether prokaryotic or eukaryotic. Let’s examine why that is so. First, we’ll consider the area and volume of a typical cell. Not all cells are spherical in shape, but most tend to approximate a sphere. You may remember from your high school geometry course that the formula for the surface area of a sphere is 4πr 2 , while the formula for its volume is 4πr 3 /3. Thus, as the radius of a cell increases, its surface area increases as the square of its radius, but its volume increases as the cube of its radius (much more rapidly). Therefore, as a cell increases in size, its surface area-to-volume ratio decreases. This same principle would apply if the cell had a cube shape ((Figure)). If the cell grows too large, the plasma membrane will not have sufficient surface area to support the rate of diffusion required for the increased volume. In other words, as a cell grows, it becomes less efficient. One way to become more efficient is to divide. Another way is to develop organelles that perform specific tasks. These adaptations lead to developing more sophisticated cells, which we call eukaryotic cells.


Prokaryotic cells are much smaller than eukaryotic cells. What advantages might small cell size confer on a cell? What advantages might large cell size have?

Section Summary

Prokaryotes are single-celled organisms of the domains Bacteria and Archaea. All prokaryotes have plasma membranes, cytoplasm, ribosomes, and DNA that is not membrane-bound. Most have peptidoglycan cell walls and many have polysaccharide capsules. Prokaryotic cells range in diameter from 0.1 to 5.0 μm.

As a cell increases in size, its surface area-to-volume ratio decreases. If the cell grows too large, the plasma membrane will not have sufficient surface area to support the rate of diffusion required for the increased volume.

Visual Connection Questions

(Figure) Prokaryotic cells are much smaller than eukaryotic cells. What advantages might small cell size confer on a cell? What advantages might large cell size have?

(Figure) Substances can diffuse more quickly through small cells. Small cells have no need for organelles and therefore do not need to expend energy getting substances across organelle membranes. Large cells have organelles that can separate cellular processes, enabling them to build molecules that are more complex.

Review Questions

Prokaryotes depend on ________ to obtain some materials and to get rid of wastes.

Bacteria that lack fimbriae are less likely to ________.

  1. adhere to cell surfaces
  2. swim through bodily fluids
  3. synthesize proteins
  4. retain the ability to divide

Which of the following organisms is a prokaryote?

Critical Thinking Questions

Antibiotics are medicines that are used to fight bacterial infections. These medicines kill prokaryotic cells without harming human cells. What part or parts of the bacterial cell do you think antibiotics target? Why?

The cell wall would be targeted by antibiotics as well as the bacteria’s ability to replicate. This would inhibit the bacteria’s ability to reproduce, and it would compromise its defense mechanisms.

Explain why not all microbes are harmful.

Some microbes are beneficial. For instance, E. coli bacteria populate the human gut and help break down fiber in the diet. Some foods such as yogurt are formed by bacteria.

Glossary


What is the specific use of a capsule in E.coli? - Biology

Chemotactic behavior

Chemotaxis, movement toward or away from chemicals, is a universal attribute of motile cells and organisms. E. coli cells swim toward amino acids (serine and aspartic acid), sugars (maltose, ribose, galactose, glucose), dipeptides, pyrimidines and electron acceptors (oxygen, nitrate, fumarate). Figure 1 shows two simple methods for assessing attractant responses by E. coli. The capillary assay relies on diffusion-generated gradients and is more quantitative, but also more laborious. Soft agar assays involve metabolism-generated chemoeffector gradients and provide a more qualitative, but expedient, measure of chemotactic ability. E. coli also swims away from potentially noxious chemicals, such as alcohols and fatty acids, but repellent responses haven't been as extensively studied.

To track chemical gradients, E. coli must solve signaling tasks common to all chemotactic behaviors: gradient detection, signal processing, and locomotor control. The chemotaxis machinery of E. coli presents a simple model for exploring the molecular mechanisms of chemosensing and signaling. However, the bizarre physical world in which E. coli lives presents unique difficulties to locomotion and calls for equally unique behavioral solutions.

Life in thermokinetic hell

The miniscule size of bacteria consigns them to a life that is dominated by viscous drag and Brownian motion (Fig. 2). Yet, despite these daunting physical constraints, E. coli cells swim at speeds of 10-20 body lengths per second. They propel themselves with locomotor organelles called flagella - very different from the flagella of eukaryotic cells - that operate much like the propellers on an ocean liner.

E. coli's optimal foraging strategy

In isotropic chemical environments, E. coli swims in a random walk pattern produced by alternating episodes of counter-clockwise (CCW) and clockwise (CW) flagellar rotation (Fig. 3, left panel). In an attractant or repellent gradient, the cells monitor chemoeffector concentration changes as they move about and use that information to modulate the probability of the next tumbling event (Fig. 3, right panel. These locomotor responses extend runs that take the cells in favorable directions (toward attractants and away from repellents), resulting in net movement toward preferred environments. Brownian motion and spontaneous tumbling episodes frequently knock the cells off course, so they must constantly assess their direction of travel with respect to the chemical gradient.

The chemotaxis signaling pathway

E. coli senses chemoeffector gradients in temporal fashion by comparing current concentrations to those encountered over the past few seconds of travel. E. coli has four transmembrane chemoreceptors, known as methyl-accepting chemotaxis proteins (MCPs), that have periplasmic ligand binding sites and conserved cytoplasmic signaling domains (Fig. 4). MCPs record the cell's recent chemical past in the form of reversible methylation of specific glutamic acid residues in the cytoplasmic signaling domain (open and filled circles in Fig. 4). Whenever current ligand occupancy state fails to coincide with the methylation record, the MCP initiates a motor control response and a feedback circuit that updates the methylation record to achieve sensory adaptation and cessation of the motor response. A fifth MCP-like protein, Aer, mediates aerotactic responses by monitoring redox changes in the electron transport chain. Aer undergoes sensory adaptation through a poorly-understood, methylation-independent mechanism.

As in many biological signaling systems, the signaling currency in the E. coli chemotaxis pathway is reversible protein phosphorylation (Fig. 5). However, the principal signaling chemistry is a bit different in prokaryotes and eukaryotes. CheA is a kinase that uses ATP to autophosphorylate at a specific histidine residue. Phospho-CheA molecules then serve as donors for autokinase reactions that transfer phosphoryl groups to specific aspartate residues in CheY and CheB. Phospho-CheY enhances CW rotation of the flagellar motors phospho-CheB has high MCP methylesterase activity. The active forms of these response regulators are short-lived because they quickly lose their phosphoryl group through spontaneous self-hydrolysis. CheZ further enhances the dephosphorylation rate of phospho-CheY to ensure rapid locomotor responses to changes in the supply of signaling phosphoryl groups to CheY.

CheW couples the autophosphorylation activity of CheA molecules to chemoreceptor control. Receptors, CheW, and CheA form stable ternary signaling complexes that modulate the influx of phosphoryl groups to the CheY and CheB proteins in response to chemoeffector stimuli.

Chemoreceptor signaling states

The signaling activities of chemoreceptors are described by a two-state model (Fig. 6). Receptor complexes in the CW signaling state activate CheA, producing high levels of phospho-CheY. Receptors in the CCW signaling state deactivate CheA, resulting in low levels of phospho-CheY. Thus, the behavior of the flagellar motors reflects the relative proportion of receptor signaling complexes in the kinase-on and kinase-off conformations. Both chemoeffector binding or release and methylation or demethylation can shift receptor signaling complexes from one state to the other. For example, attractant ligands drive receptors toward the kinase-off state subsequent addition of methyl groups shifts receptors toward the kinase-on state, reestablishing the steady-state (adapted) balance between the two states and restoring random walk movements.


Prokaryotic Recombination

Genetic variation within prokaryotic organisms is accomplished through recombination. In recombination, genes from one prokaryote are incorporated into the genome of another prokaryote.

Recombination is accomplished in bacterial reproduction by the processes of conjugation, transformation, or transduction.