Gram-negative bacteria#Taxonomy

File:Gram negative cell wall.svg structure]]

File:Gram-Cell-wall.svg and -negative bacteria are differentiated chiefly by their cell wall structure]]

Conventional gram-negative (LPS-diderm) bacteria display {{nowrap|the following characteristics}}:{{cn|date=July 2022}}

• An inner cell membrane is present (cytoplasmic)
• A thin peptidoglycan layer is present (this is much thicker in gram-positive bacteria)
• Has outer membrane containing lipopolysaccharides (LPS, which consists of lipid A, core polysaccharide, and O antigen) in its outer leaflet and phospholipids in the inner leaflet
• Porins exist in the outer membrane, which act like pores for particular molecules
• Between the outer membrane and the cytoplasmic membrane there is a space filled with a concentrated gel-like substance called periplasm
• The S-layer is directly attached to the outer membrane rather than to the peptidoglycan
• If present, flagella have four supporting rings instead of two
• Teichoic acids or lipoteichoic acids are absent
• Lipoproteins are attached to the polysaccharide backbone
• Some contain Braun's lipoprotein, which serves as a link between the outer membrane and the peptidoglycan chain by a covalent bond
• Most, with few exceptions, do not form spores

However, the LPS-diderm group (corresponding to kingdom Pseudomonadati, formerly 'Hydrobacteria') is not the only type of bacteria that stain negative.{{cite journal |last1=Göker |first1=Markus |last2=Oren |first2=Aharon |title=Valid publication of names of two domains and seven kingdoms of prokaryotes |journal=International Journal of Systematic and Evolutionary Microbiology |date=22 January 2024 |volume=74 |issue=1 |doi=10.1099/ijsem.0.006242|pmid=38252124 }} Mycobacterium (or rather most of Mycobacteriales), which does not belong in the group, have independently evolved an outer cell membrane, with a cell wall made of mycolic acid.{{cite journal |last1=Gupta |first1=Radhey S. |title=Commentary: Genome-Based Taxonomic Classification of the Phylum Actinobacteria |journal=Frontiers in Microbiology |date=22 February 2019 |volume=10 |page=206 |doi=10.3389/fmicb.2019.00206 |quote=Mycolic acids are important constituents of the cell envelopes of most members.|doi-access=free |pmid=30853945 |pmc=6395429 }} This gives it very different structure and features.{{cite journal|vauthors=Marchandin H, Teyssier C, Campos J, Jean-Pierre H, Roger F, Gay B, Carlier JP, Jumas-Bilak E |title=Negativicoccus succinicivorans gen. nov., sp. nov., isolated from human clinical samples, emended description of the family Veillonellaceae and description of Negativicutes classis nov., Selenomonadales ord. nov. and Acidaminococcaceae fam. nov. in the bacterial phylum Firmicutes |journal=Int. J. Syst. Evol. Microbiol. |volume=60 |issue=Pt 6 |pages=1271–9 |date=June 2010 |pmid=19667386 |doi=10.1099/ijs.0.013102-0|doi-access=free }}

Along with cell shape, Gram staining is a rapid diagnostic tool and once was used to group species at the subdivision of Bacteria.

Historically, the kingdom Monera was divided into four divisions based on Gram staining: Firmicutes (+), Gracillicutes (−), Mollicutes (0) and Mendocutes (var.).{{cite journal |last1=Gibbons |first1=N. E. |last2=Murray |first2=R. G. E. |title=Proposals Concerning the Higher Taxa of Bacteria |journal=International Journal of Systematic Bacteriology |year=1978 |volume=28 |issue=1 |pages=1–6 |doi=10.1099/00207713-28-1-1 |doi-access=free }}

Since 1987, the monophyly of the gram-negative bacteria has been disproven with molecular studies.{{cite journal | author = Woese CR | title = Bacterial evolution | journal = Microbiol. Rev. | volume = 51 | issue = 2 | pages = 221–71 |date=June 1987 | pmid = 2439888 | pmc = 373105 | doi = 10.1128/MMBR.51.2.221-271.1987}}

Current knowledge divides the gram-negatives into two large groups and some straddlers. The more "conventional" gram-negatives with an LPS outer membrane do share a common ancestor and are grouped in kingdom Pseudomonadati. The less conventional ones are, as mentioned above, the order Mycobacteriales, have a mycolic acid cell wall and an outer membrane. The kingdom and the order are each monophyletic (or rather, not holyphyletic), but the "LPS-diderm" and "mycolic-diderm" groups are not, because some bacteria in the kingdom and the order do not, in fact, stain gram negative. They will be discussed in the next section.

{{Main|Bacterial taxonomy}}

{{Further|Bacteria#Classification and identification}}

{{Technical|section|date=March 2014}}

{{Split section|Gram stain |discuss=Gram stain#Move Taxonomy sections here |date=November 2023}}

Bacteria are traditionally classified based on their Gram-staining response into the gram-positive and gram-negative bacteria. Having just one membrane, the gram-positive bacteria are also known as monoderm bacteria, while gram-negative bacteria, having two membranes, are also known as diderm bacteria. It was traditionally thought that the groups represent lineages, i.e., the extra membrane only evolved once, such that gram-negative bacteria are more closely related to one another than to any gram-positive bacteria. While this is often true, the classification system breaks down in some cases, with lineage groupings not matching the staining result.{{cite journal|last=Gupta |first=RS |title=Protein phylogenies and signature sequences: A reappraisal of evolutionary relationships among archaebacteria, eubacteria, and eukaryotes |journal=Microbiol. Mol. Biol. Rev. |volume=62 |issue=4 |pages=1435–91 |date=December 1998 |pmid=9841678 |pmc=98952 |doi=10.1128/MMBR.62.4.1435-1491.1998}}{{cite journal | author = Gupta RS | title = The natural evolutionary relationships among prokaryotes | journal = Crit. Rev. Microbiol. | volume = 26 | issue = 2 | pages = 111–31 | year = 2000 | pmid = 10890353 | doi = 10.1080/10408410091154219 | url = http://www.life.illinois.edu/govindjee/Part2/15_Gupta.pdf | citeseerx = 10.1.1.496.1356 | s2cid = 30541897 | access-date = 2017-10-24 | archive-date = 2018-07-20 | archive-url = https://web.archive.org/web/20180720153253/http://www.life.illinois.edu/govindjee/Part2/15_Gupta.pdf | url-status = live }}{{cite journal |vauthors=Desvaux M, Hébraud M, Talon R, Henderson IR | title = Secretion and subcellular localizations of bacterial proteins: a semantic awareness issue | journal = Trends Microbiol. | volume = 17 | issue = 4 | pages = 139–45 |date=April 2009 | pmid = 19299134 | doi = 10.1016/j.tim.2009.01.004 }}{{cite journal | author = Sutcliffe IC | title = A phylum level perspective on bacterial cell envelope architecture | journal = Trends Microbiol. | volume = 18 | issue = 10 | pages = 464–70 |date=October 2010 | pmid = 20637628 | doi = 10.1016/j.tim.2010.06.005 }} Thus, Gram staining cannot be reliably used to assess familial relationships of bacteria. Nevertheless, staining often gives reliable information about the composition of the cell membrane, distinguishing between the presence or absence of an outer lipid membrane.{{cite journal | author = Gupta RS | title = What are archaebacteria: life's third domain or monoderm prokaryotes related to gram-positive bacteria? A new proposal for the classification of prokaryotic organisms | journal = Mol. Microbiol. | volume = 29 | issue = 3 | pages = 695–707 |date=August 1998 | pmid = 9723910 | doi = 10.1046/j.1365-2958.1998.00978.x | doi-access = free }}

Of these two structurally distinct groups of prokaryotic organisms, monoderm prokaryotes are thought to be ancestral. Based upon a number of different observations, including that the gram-positive bacteria are the most sensitive to antibiotics and that the gram-negative bacteria are, in general, resistant to antibiotics, it has been proposed that the outer cell membrane in gram-negative bacteria (diderms) evolved as a protective mechanism against antibiotic selection pressure.{{cite journal | author = Gupta RS | title = Origin of diderm (gram-negative) bacteria: antibiotic selection pressure rather than endosymbiosis likely led to the evolution of bacterial cells with two membranes | journal = Antonie van Leeuwenhoek | volume = 100 | issue = 2 | pages = 171–82 |date=August 2011 | pmid = 21717204 | pmc = 3133647 | doi = 10.1007/s10482-011-9616-8 }} Some bacteria such as Deinococcus, which stain gram-positive due to the presence of a thick peptidoglycan layer, but also possess an outer cell membrane are suggested as intermediates in the transition between monoderm (gram-positive) and diderm (gram-negative) bacteria.

The conventional LPS-diderm group of gram-negative bacteria (e.g., Pseudomonadota, Aquificota, Chlamydiota, Bacteroidota, Chlorobiota, "Cyanobacteria", Fibrobacterota, Verrucomicrobiota, Planctomycetota, Spirochaetota, Acidobacteriota) are uniquely identified by a few conserved signature indel (CSI) in the HSP60 (GroEL) protein. The presence of this CSI in all sequenced species of conventional lipopolysaccharide-containing gram-negative bacterial phyla provides evidence that these phyla of bacteria form a monophyletic clade and that no loss of the outer membrane from any species from this group has occurred. They have accordingly been assigned a kingdom Pseudomonadati (formerly "Hydrobacteria").

The difficulty lies in the other taxa that also have a diderm structure.

• The first group is paraphyletic. It includes a number of taxa (including Negativicutes, Fusobacteriota, Synergistota, and Elusimicrobiota) that are either part of the phylum Bacillota (a monoderm group) or branches in its proximity. They lack the GroEL CSI signature, which is proof that they do not belong in the former group. Some members are likely monoderm, just with a very thin layer of LPS to not appear on the stain. Others have more convoluted structures.{{cite journal |last1=Choi |first1=JK |last2=Poudel |first2=S |last3=Yee |first3=N |last4=Goff |first4=JL |title=Deeply branching Bacillota species exhibit atypical Gram-negative staining. |journal=Microbiology Spectrum |date=3 October 2024 |volume=12 |issue=10 |pages=e0073224 |doi=10.1128/spectrum.00732-24 |pmid=39162559|pmc=11448272 }}
• The second group are the clinically-relevant Mycobacterium, expanding to most of its encompassing order of Mycobacteriales. They do not have the CSI, and their cell wall is made of a different substance: mycolic acid.

The proteobacteria are a major superphylum of gram-negative bacteria, including E. coli, Salmonella, Shigella, and other Enterobacteriaceae, Pseudomonas, Moraxella, Helicobacter, Stenotrophomonas, Bdellovibrio, acetic acid bacteria, Legionella etc. Other notable groups of gram-negative bacteria include the cyanobacteria, spirochaetes, and green sulfur bacteria.{{Citation |last=Castenholz |first=Richard W. |title=General Characteristics of the Cyanobacteria |date=2015 |work=Bergey's Manual of Systematics of Archaea and Bacteria |pages=1–23 |url=https://onlinelibrary.wiley.com/doi/10.1002/9781118960608.cbm00019 |access-date=2025-01-25 |publisher=John Wiley & Sons, Ltd |language=en |doi=10.1002/9781118960608.cbm00019 |isbn=978-1-118-96060-8}}{{Citation |last=Cole |first=John R. |title=5 - Spirochetes |date=1990-01-01 |work=Diagnostic Procedure in Veterinary Bacteriology and Mycology (Fifth Edition) |pages=41–60 |editor-last=Carter |editor-first=G. R. |url=https://linkinghub.elsevier.com/retrieve/pii/B9780121617752500098 |access-date=2025-01-25 |place=San Diego |publisher=Academic Press |doi=10.1016/b978-0-12-161775-2.50009-8 |isbn=978-0-12-161775-2 |editor2-last=Cole |editor2-first=John R.}}{{Cite book |last=Schmidt |first=Thomas M. |title=Encyclopedia of Microbiology |date=2019 |publisher=Elsevier Science & Technology |isbn=978-0-12-811737-8 |edition=4th |location=San Diego |pages=527–537}}

Medically-relevant gram-negative diplococci include the four types that cause a sexually transmitted disease (Neisseria gonorrhoeae{{cite journal |last1=Yeshanew |first1=Addisu Gize |last2=Geremew |first2=Rozina Ambachew |date=2018-07-17 |title=MNeisseria Gonorrhoae and their antimicrobial susceptibility patterns among symptomatic patients from Gondar town, north West Ethiopia |journal=Antimicrobial Resistance and Infection Control |volume=7 |issue=85 |page=85 |doi=10.1186/s13756-018-0376-3 |doi-access=free |pmid=30026943 |pmc=6050735 }}), a meningitis (Neisseria meningitidis{{cite web | title=Vaccine Preventable Diseases Surveillance Manual | website=CDC | date=April 3, 2018 | url=https://www.cdc.gov/vaccines/pubs/surv-manual/chpt08-mening.html | access-date=January 26, 2024}}), and respiratory symptoms (Moraxella catarrhalis,{{cite journal |last1=Verduin |first1=Cees M. |last2=Hol |first2=Cees |last3=Fleer |first3=Andre |last4=van Dijk |first4=Hans |last5=van Belkum |first5=Alex |date=January 2002 |title=Moraxella catarrhalis: from Emerging to Established Pathogen |journal=Clinical Microbiology Reviews |volume=15 |issue=1 |pages=125–144 |doi=10.1128/CMR.15.1.125-144.2002 |pmid=11781271 |pmc=118065 }} A coccobacillus Haemophilus influenzae is another medically relevant coccal type.{{cite web | title=For Clinicians: Haemophilus influenzae | website=CDC | date=February 13, 2018 | url=https://www.cdc.gov/hi-disease/clinicians.html#:~:text=Haemophilus%20influenzae%20is%20a%20pleomorphic,that%20have%20distinct%20capsular%20polysaccharides. | access-date=January 26, 2024}}

Medically relevant gram-negative bacilli include a multitude of species. Some of them cause primarily respiratory problems (Klebsiella pneumoniae, Legionella pneumophila, Pseudomonas aeruginosa), primarily urinary problems (Escherichia coli, Proteus mirabilis, Enterobacter cloacae, Serratia marcescens), and primarily gastrointestinal problems (Helicobacter pylori, Salmonella enteritidis, Salmonella typhi).{{cn|date=November 2023}}

Gram-negative bacteria associated with hospital-acquired infections include Acinetobacter baumannii, which cause bacteremia, secondary meningitis, and ventilator-associated pneumonia in hospital intensive-care units.

Transformation is one of three processes for horizontal gene transfer, in which exogenous genetic material passes from one bacterium to another, the other two being conjugation (transfer of genetic material between two bacterial cells in direct contact) and transduction (injection of foreign DNA by a bacteriophage virus into the host bacterium).{{cite journal |vauthors=Johnston C, Martin B, Fichant G, Polard P, Claverys JP |title=Bacterial transformation: distribution, shared mechanisms and divergent control |journal=Nat. Rev. Microbiol. |volume=12 |issue=3 |pages=181–96 |year=2014 |pmid=24509783 |doi=10.1038/nrmicro3199 |s2cid=23559881 }}{{cite journal |vauthors=Korotetskiy I, Shilov S, Kuznetsova T, Kerimzhanova B, Korotetskaya N, Ivanova L, Zubenko N, Parenova R, Reva O |title=Analysis of Whole-Genome Sequences of Pathogenic Gram-Positive and Gram-Negative Isolates from the Same Hospital Environment to Investigate Common Evolutionary Trends Associated with Horizontal Gene Exchange, Mutations and DNA Methylation Patterning |journal=Microorganisms |volume=11 |issue=2 |year=2023 |page=323 |pmid=36838287 |doi=10.3390/microorganisms11020323 |doi-access=free |pmc=9961978 }} In transformation, the genetic material passes through the intervening medium, and uptake is completely dependent on the recipient bacterium.

As of 2014 about 80 species of bacteria were known to be capable of transformation, about evenly divided between gram-positive and gram-negative bacteria; the number might be an overestimate since several of the reports are supported by single papers. Transformation has been studied in medically important gram-negative bacteria species such as Helicobacter pylori, Legionella pneumophila, Neisseria meningitidis, Neisseria gonorrhoeae, Haemophilus influenzae and Vibrio cholerae.{{cite journal |vauthors=Seitz P, Blokesch M |title=Cues and regulatory pathways involved in natural competence and transformation in pathogenic and environmental Gram-negative bacteria |journal=FEMS Microbiol. Rev. |volume=37 |issue=3 |pages=336–63 |year=2013 |pmid=22928673 |doi=10.1111/j.1574-6976.2012.00353.x |doi-access=free }} It has also been studied in gram-negative species found in soil such as Pseudomonas stutzeri, Acinetobacter baylyi, and gram-negative plant pathogens such as Ralstonia solanacearum and Xylella fastidiosa.

File:Diagnostic algorithm of possible bacterial infection.png and/or multitarget assay.]]

One of the several unique characteristics of gram-negative bacteria is the structure of the bacterial outer membrane. The outer leaflet of this membrane contains lipopolysaccharide (LPS), whose lipid A portion acts as an endotoxin. If gram-negative bacteria enter the circulatory system, LPS can trigger an innate immune response, activating the immune system and producing cytokines (hormonal regulators). This leads to inflammation and can cause a toxic reaction, resulting in fever, an increased respiratory rate, and low blood pressure. That is why some infections with gram-negative bacteria can lead to life-threatening septic shock.

The outer membrane protects the bacteria from several antibiotics, dyes, and detergents that would normally damage either the inner membrane or the cell wall (made of peptidoglycan). The outer membrane provides these bacteria with resistance to lysozyme and penicillin. The periplasmic space (space between the two cell membranes) also contains enzymes which break down or modify antibiotics. Drugs commonly used to treat gram negative infections include amino, carboxy and ureido penicillins (ampicillin, amoxicillin, pipercillin, ticarcillin). These drugs may be combined with beta-lactamase inhibitors to combat the presence of enzymes that can digest these drugs (known as beta-lactamases) in the peri-plasmic space. Other classes of drugs that have gram negative spectrum include cephalosporins, monobactams (aztreonam), aminoglycosides, quinolones, macrolides, chloramphenicol, folate antagonists, and carbapenems.{{cite web|last=Glück|first=Thomas|url=https://www.jwatch.org/id200312120000007/2003/12/12/gram-negative-bacteria-and-broad-spectrum|title=Gram-Negative Bacteria and Broad-Spectrum Antibiotics: Good News Except for Fluoroquinolones|publisher=NEJM Journal Watch|website=www.jwatch.org|date=12 December 2003|access-date=10 March 2018|archive-date=10 March 2018|archive-url=https://web.archive.org/web/20180310074653/https://www.jwatch.org/id200312120000007/2003/12/12/gram-negative-bacteria-and-broad-spectrum|url-status=live}}

The adjectives gram-positive and gram-negative derive from the surname of Hans Christian Gram, a Danish bacteriologist; as eponymous adjectives, their initial letter can be either capital G or lower-case g, depending on which style guide (e.g., that of the CDC), if any, governs the document being written.{{cite web |publisher=Centers for Disease Control and Prevention |title=Preferred Usage - Emerging Infectious Disease journal - CDC |url=https://wwwnc.cdc.gov/eid/page/preferred-usage |work=CDC.gov |access-date=2018-03-04 |archive-date=2018-01-29 |archive-url=https://web.archive.org/web/20180129223909/https://wwwnc.cdc.gov/eid/page/preferred-usage |url-status=live }} This is further explained at Gram staining § Orthographic note.

• Autochaperone
• Gram-variable and gram-indeterminate bacteria
• OMPdb (2011)
• Outer membrane receptor

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• [https://web.archive.org/web/20081222045655/http://opm.phar.umich.edu:80/localization.php?localization=Bacterial%20gram-negative%20inner%20membrane 3D structures of proteins from inner membranes of Ellie Wyithe's gram-negative bacteria]

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Category:Staining

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