Gram-negative
Gram-negative bacteria are bacteria that do not retain the crystal violet stain used in the Gram staining method of bacterial differentiation. They are characterized by their cell envelopes, which are composed of a thin peptidoglycan cell wa ...
bacteria
Bacteria (; singular: bacterium) are ubiquitous, mostly free-living organisms often consisting of one Cell (biology), biological cell. They constitute a large domain (biology), domain of prokaryotic microorganisms. Typically a few micrometr ...
in the genus ''
Shigella
''Shigella'' is a genus of bacteria that is Gram-negative, facultative anaerobic, non-spore-forming, nonmotile, rod-shaped, and genetically closely related to '' E. coli''. The genus is named after Kiyoshi Shiga, who first discovered it in 1 ...
'' that can cause diarrhea in humans. Several different serogroups of ''Shigella'' are described; ''S. flexneri'' belongs to group ''B''. ''S. flexneri'' infections can usually be treated with antibiotics, although some strains have become resistant. Less severe cases are not usually treated because they become more resistant in the future. Shigella are closely related to ''Escherichia coli'', but can be differentiated from ''E.coli'' based on pathogenicity, physiology (failure to ferment lactose or decarboxylate lysine) and serology.
Discovery
The species was named after the American physician
Simon Flexner
Simon Flexner, M.D. (March 25, 1863 in Louisville, Kentucky – May 2, 1946) was a physician, scientist, administrator, and professor of experimental pathology at the University of Pennsylvania (1899–1903). He served as the first director ...
; the genus Shigella is named after Japanese physician Kiyoshi Shiga, who researched the cause of dysentery. Shiga entered the Tokyo Imperial University School of Medicine in 1892, during which he attended a lecture by Dr. Shibasaburo Kitasato. Shiga was impressed by Dr. Kitasato's intellect and confidence, so after graduating, he went to work for him as a research assistant at Institute for Infectious Diseases. In 1897, Shiga focused his efforts on what the Japanese referred to as a "Sekiri" (dysentery) outbreak. These epidemics were detrimental to the Japanese people and occurred often in the late 19th century. The 1897 ''sekiri'' epidemic affected >91,000, with a mortality rate of >20%. Shiga studied 32 dysentery patients and used
Shiga Toxin
Shiga toxins are a family of related toxins with two major groups, Stx1 and Stx2, expressed by genes considered to be part of the genome of lambdoid prophages. The toxins are named after Kiyoshi Shiga, who first described the bacterial or ...
, and worked tirelessly to create a vaccine for the disease.
Characterization
Morphology
''Shigella flexneri'' is a rod shaped, nonflagellar bacterium that relies on actin-based motility. It produces the protein actin in a swift and continuous fashion to propel itself forward within and between the host’s cells. This bacterium is gram-negative, non-spore forming ''Shigella'' from serogroup B. There are 6 serotypes within this serogroup.
Serotype
''Shigella flexneri'' belongs to group B (i.e. agglutinate with B antisera) which further subclassified by six type-specific and four group-specific antisera. Until now at least 23 different subserotypes have been identified and reported. PCR based molecular serotyping technique are now available targeting wzx1-5 (All except serotype 6) and gtr genes or wzx6 (Only serotype 6).
Invasion
''Shigella flexneri'' is an intracellular bacterium that infects the epithelial lining of the mammalian intestinal tract. This bacterium is acid tolerant and can survive conditions of pH 2. Thus, it is able to enter the mouth of its host and survive passage through the stomach to the colon. Once inside of the colon, ''S. flexneri'' can penetrate the epithelium in three ways: 1) The bacterium can alter the tight junctions between the epithelial cells, allowing it to cross into the sub-mucosa. 2) It can penetrate the highly endocytic M cells that are dispersed in the epithelial layer and cross into the sub-mucosa. 3) After reaching the sub-mucosa, the bacteria can be phagocytosed by macrophages and induce apoptosis, cell death. This releases
cytokine
Cytokines are a broad and loose category of small proteins (~5–25 kDa) important in cell signaling. Cytokines are peptides and cannot cross the lipid bilayer of cells to enter the cytoplasm. Cytokines have been shown to be involved in autocrin ...
s that recruit polymorphonuclear cells (PMN) to the sub-mucosa. ''S. flexneri'' still in the lumen of the colon traverse the epithelial lining as the PMNs cross into the infected area. The influx of PMN cells across the epithelial layer in response to Shigella disrupts the integrity of the epithelium allowing lumenal bacteria to cross into the sub-mucosa in an M-cell independent mechanism. ''S. flexneri'' uses these three methods to reach the sub-mucosa to penetrate the epilithelial cells from the basolateral side. The bacterium has four known invasion plasmid antigens: IpaA, IpaB, IpaC, and IpaD. When ''S. flexneri'' makes contact with the basolateral side of an epithelial cell, IpaC and IpaB are fused together to make a pore in the epithelial cell membrane. It then uses a type-III secretion system (T3SS) to insert the other Ipa proteins into the cytoplasm of the epithelial cell. ''S. flexneri'' can pass to neighboring epithelial cells by using its own outer membrane protein, IcsA, to activate the host's actin assembly machinery. The IcsA protein is first localized to one pole of the bacterium where it will then bind with the host's protein, Neural Wiskott-Aldrich Syndrome Protein (N-WASP). This IcsA/N-WASP complex then activates the Actin-related protein (Arp) 2/3 Complex. Arp 2/3 Complex is the protein responsible for rapidly initiating actin polymerization and propelling the bacteria forward. When ''S. flexneri'' reaches the adjoining membrane, it creates a protrusion into the neighboring cell's cytoplasm. The bacteria becomes surrounded by two layers of cellular membrane. It then uses another IpaBC complex to make a pore and enter the next cell. VacJ is a protein that is also needed by ''S. flexneri'' to exit the protrusion. Its exact function is still being studied but it is known that intercellular spread is greatly impaired without it. Bacterial replication within the epithelial cell is detrimental to the cell but it is proposed that epithelial cell death is largely due to the host’s own inflammatory response.
Genetics
The genome of ''S. flexneri'' and ''Escherichia coli'' are nearly indistinguishable at the species level. ''S. flexneri'' has a circular chromosome with 4,599,354 base pairs. It is smaller than that of ''E. coli'' but the genes are similar. ''S. flexneri'' has about 4,084 known genes in the genome. The extensive similarity between ''E. coli'' and ''S. flexneri'' is proposed to be due to horizontal transfer. All of the genes needed for ''S. flexneri'' to invade the epithelial lining of the colon are found on a virulence plasmid called pINV. The genome of pINV is highly conserved between subspecies of ''S. flexneri''. ''S. flexneri'' also has two other small multicopy plasmids, but some strains of ''S. flexneri'' have more plasmids that are suspected to confer antibiotic resistance. Some strains of ''S. flexneri'' have resistance to the antibiotics streptomycin, ampicillin, or trimethoprim. It has been found that chloramphenicol, nalidixic acid, and gentamicin are still effective antibiotics for some strains.
Metabolism
''Shigella flexneri'' is a
heterotroph
A heterotroph (; ) is an organism that cannot produce its own food, instead taking nutrition from other sources of organic carbon, mainly plant or animal matter. In the food chain, heterotrophs are primary, secondary and tertiary consumers, but ...
. It utilizes the Embden-Meyerhof-Parnas (EMP), Entner-Doudoroff (ED), or pentose phosphate pathway (PPP) to metabolize sugars. The products of these pathways then feed into the Citric Acid Cycle (TCA). ''S. flexneri'' can metabolize glucose and pyruvate. Supplemented pyruvate allows for the most growth and is believed to be the preferred carbon source. Pyruvate could be supplied by the cell's own metabolism or taken from the host cell. ''S. flexneri'' is a
facultative anaerobe
A facultative anaerobic organism is an organism that makes ATP by aerobic respiration if oxygen is present, but is capable of switching to fermentation if oxygen is absent.
Some examples of facultatively anaerobic bacteria are '' Staphylococc ...
that is able to perform mixed-acid fermentation of pyruvate. ''S. flexneri'' is unable to ferment lactose. This bacterium grows optimally at 37°C but can grow in temperatures as low as 30°C.
Small RNA
Bacterial small RNAs play important roles in many cellular processes. RnaG and RyhB sRNAs have been well studied in ''S. flexneri''. Ssr1 sRNA, which could play role in resistance to acidic stress and regulation of virulence was shown to exist only in ''Shigella''.
Ecology
Infectious cycle
''Shigella flexneri'' contains a virulence plasmid that codes for three virulence factors: a type-3 secretion system (T3SS), invasion plasmid antigen proteins (IPA proteins), and IcsA (used for cell-to-cell spread).
Upon infection, ''S. flexneri'' injects the host cell cytoplasm with ipa proteins using the T3SS—a needle-and-syringe-like apparatus common to many Gram-negative pathogens. These ipa proteins induce "membrane ruffling" by the host cell. Membrane ruffling creates membrane pockets which capture and engulf the bacteria. Once inside, ''S. flexneri'' uses host cell actin for propulsion to move directly from cell to cell using a cellular mechanism known as paracytophagy, similarly to the bacterial pathogen ''
Listeria monocytogenes
''Listeria monocytogenes'' is the species of pathogenic bacteria that causes the infection listeriosis. It is a facultative anaerobic bacterium, capable of surviving in the presence or absence of oxygen. It can grow and reproduce inside the host ...
''.
''Shigella flexneri'' is able to inhibit the acute inflammatory response in the initial stage of infection by using an effector protein, OspI, which is encoded by ''ORF169b'' on the ''Shigella'' large plasmid and secreted by the type III secretion system. It dampens the inflammatory response during bacterial invasion by suppressing the
TNF-α
Tumor necrosis factor (TNF, cachexin, or cachectin; formerly known as tumor necrosis factor alpha or TNF-α) is an adipokine and a cytokine. TNF is a member of the TNF superfamily, which consists of various transmembrane proteins with a homolog ...
-receptor-associated factor 6 (
TRAF6
TRAF6 is a TRAF human protein.
Function
The protein encoded by this gene is a member of the TNF receptor associated factor (TRAF) protein family. TRAF proteins are associated with, and mediate signal transduction from members of the TNF recep ...
)-mediated signalling pathway. OspI has glutamine deamidase activity, and is able to selectively deaminate
glutamine
Glutamine (symbol Gln or Q) is an α-amino acid that is used in the biosynthesis of proteins. Its side chain is similar to that of glutamic acid, except the carboxylic acid group is replaced by an amide. It is classified as a charge-neutral ...
at position 100 in UBC13 to glutamate, and this results in a failure of the E2 ubiquitin-conjugating activity which is required for TRAF6 activation.