Editing Virology (section)

==Detecting viruses==
[[File:Jeol Transmission and scanning EM.jpg|thumb|right|An electron microscope]]
There are several approaches to detecting viruses and these include the detection of virus particles (virions) or their [[antigens]] or nucleic acids and infectivity assays.

===Electron microscopy===
[[File:Gastroenteritis viruses.jpg|right|thumb|Electron micrographs of viruses. A, rotavirus; B, adenovirus; C, norovirus; and D, astrovirus.]]
Viruses were seen for the first time in the 1930s when electron microscopes were invented. These microscopes use beams of [[electron]]s instead of light, which have a much shorter wavelength and can detect objects that cannot be seen using light microscopes. The highest magnification obtainable by electron microscopes is up to 10,000,000 times<ref name =Payne>Payne S. Methods to Study Viruses. Viruses. 2017;37-52. doi:10.1016/B978-0-12-803109-4.00004-0</ref> whereas for light microscopes it is around 1,500 times.<ref>{{cite web|title=Magnification - Microscopy, size and magnification (CCEA) - GCSE Biology (Single Science) Revision - CCEA|url=https://www.bbc.co.uk/bitesize/guides/z3vypbk/revision/3|access-date=2023-01-02|website=BBC Bitesize|language=en-GB}}</ref>

Virologists often use [[negative staining#Transmission electron microscopy|negative staining]] to help visualise viruses. In this procedure, the viruses are suspended in a solution of metal salts such as uranium acetate. The atoms of metal are opaque to electrons and the viruses are seen as suspended in a dark background of metal atoms.<ref name =Payne/> This technique has been in use since the 1950s.<ref name="pmid13804200">{{cite journal |vauthors=Brenner S, Horne RW |title=A negative staining method for high resolution electron microscopy of viruses |journal=Biochimica et Biophysica Acta |volume=34 |issue= |pages=103–10 |date=July 1959 |pmid=13804200 |doi=10.1016/0006-3002(59)90237-9}}</ref> Many viruses were discovered using this technique and negative staining electron microscopy is still a valuable weapon in a virologist's arsenal.<ref name="pmid19822888">{{cite journal |vauthors=Goldsmith CS, Miller SE |title=Modern uses of electron microscopy for detection of viruses |journal=Clinical Microbiology Reviews |volume=22 |issue=4 |pages=552–63 |date=October 2009 |pmid=19822888 |pmc=2772359 |doi=10.1128/CMR.00027-09}}</ref>

Traditional electron microscopy has disadvantages in that viruses are damaged by drying in the high vacuum inside the electron microscope and the electron beam itself is destructive.<ref name=Payne/> In [[cryogenic electron microscopy]] the structure of viruses is preserved by embedding them in an environment of [[vitreous water]].<ref>{{cite journal|last1=Tivol|first1=William F.|last2=Briegel|first2=Ariane|last3=Jensen|first3=Grant J.|date=October 2008|title=An Improved Cryogen for Plunge Freezing|journal=Microscopy and Microanalysis|language=en|volume=14|issue=5|pages=375–379|doi=10.1017/S1431927608080781|issn=1431-9276|pmc=3058946|pmid=18793481|bibcode=2008MiMic..14..375T}}</ref> This allows the determination of biomolecular structures at near-atomic resolution,<ref>{{cite journal | vauthors = Cheng Y, Grigorieff N, Penczek PA, Walz T | title = A primer to single-particle cryo-electron microscopy | journal = Cell | volume = 161 | issue = 3 | pages = 438–449 | date = April 2015 | pmid = 25910204 | pmc = 4409659 | doi = 10.1016/j.cell.2015.03.050 }}</ref> and has attracted wide attention to the approach as an alternative to [[X-ray crystallography]] or [[NMR spectroscopy]] for the determination of the structure of viruses.<ref name="Stoddart">{{cite journal |last1=Stoddart |first1=Charlotte |title=Structural biology: How proteins got their close-up |journal=Knowable Magazine |date=1 March 2022 |doi=10.1146/knowable-022822-1|doi-access=free |url=https://knowablemagazine.org/article/living-world/2022/structural-biology-how-proteins-got-their-closeup |access-date=25 March 2022}}</ref>
[[File:Rotavirus Reconstruction.jpg|right|thumb|Cryoelectron micrograph of a rotavirus]]

===Growth in cultures===
Viruses are obligate intracellular parasites and because they only reproduce inside the living cells of a host these cells are needed to grow them in the laboratory. For viruses that infect animals (usually called "animal viruses") cells grown in laboratory [[cell culture]]s are used. In the past, fertile hens' eggs were used and the viruses were grown on the membranes surrounding the embryo. This method is still used in the manufacture of some vaccines. For the viruses that infect bacteria, the [[bacteriophages]], the bacteria growing in test tubes can be used directly. For plant viruses, the natural host plants can be used or, particularly when the infection is not obvious, so-called indicator plants, which show signs of infection more clearly.<ref name="pmid19653216">{{cite journal |vauthors=Liu JZ, Richerson K, Nelson RS |title=Growth conditions for plant virus-host studies |journal=Current Protocols in Microbiology |volume=Chapter 16 |issue= |pages=Unit16A.1 |date=August 2009 |pmid=19653216 |doi=10.1002/9780471729259.mc16a01s14 |s2cid=41236532 |url=}}</ref><ref name="pmid35050076">{{cite journal |vauthors=Valmonte-Cortes GR, Lilly ST, Pearson MN, [[Colleen Higgins|Higgins CM]], MacDiarmid RM |date=January 2022 |title=The Potential of Molecular Indicators of Plant Virus Infection: Are Plants Able to Tell Us They Are Infected? |url= |journal=Plants |volume=11 |issue=2 |page=188 |doi=10.3390/plants11020188 |pmc=8777591 |pmid=35050076 |doi-access=free}}</ref>
[[File:CPE rounding.jpg|right|thumb|Cytopathic effect of herpes simplex virus. The infected cells have become round and balloon-like.]]
Viruses that have grown in cell cultures can be indirectly detected by the detrimental effect they have on the host cell. These [[cytopathic effect]]s are often characteristic of the type of virus. For instance, [[herpes simplex virus]]es produce a characteristic "ballooning" of the cells, typically human [[fibroblast]]s. Some viruses, such as [[mumps virus]] cause red blood cells from chickens to firmly attach to the infected cells. This is called "haemadsorption" or "hemadsorption". Some viruses produce localised "lesions" in cell layers called [[Viral plaque|plaques]], which are useful in quantitation assays and in identifying the species of virus by [[plaque forming units|plaque reduction assays]].<ref name="pmid24899440">{{cite book |vauthors=Gauger PC, Vincent AL |chapter=Serum Virus Neutralization Assay for Detection and Quantitation of Serum-Neutralizing Antibodies to Influenza a Virus in Swine |title=Animal Influenza Virus |series=Methods in Molecular Biology |volume=1161 |pages=313–24 |date=2014 |pmid=24899440 |doi=10.1007/978-1-4939-0758-8_26 |isbn=978-1-4939-0757-1 |chapter-url=}}</ref><ref name="pmid32367358">{{cite book |vauthors=Dimitrova K, Mendoza EJ, Mueller N, Wood H |title=Zika Virus |chapter=A Plaque Reduction Neutralization Test for the Detection of ZIKV-Specific Antibodies |series=Methods in Molecular Biology |volume=2142 |pages=59–71 |date=2020 |pmid=32367358 |doi=10.1007/978-1-0716-0581-3_5 |isbn=978-1-0716-0580-6 |s2cid=218504421 |chapter-url=}}</ref>

Viruses growing in cell cultures are used to measure their susceptibility to validated and novel [[antiviral drug]]s.<ref name="pmid32056049">{{cite journal |vauthors=Lampejo T |title=Influenza and antiviral resistance: an overview |journal=European Journal of Clinical Microbiology & Infectious Diseases |volume=39 |issue=7 |pages=1201–1208 |date=July 2020 |pmid=32056049 |pmc=7223162 |doi=10.1007/s10096-020-03840-9 |url=}}</ref>

===Serology===
Viruses are [[antigens]] that induce the production of [[antibodies]] and these antibodies can be used in laboratories to study viruses. Related viruses often react with each other's antibodies and some viruses can be named based on the antibodies they react with. The use of the antibodies which were once exclusively derived from the serum (blood fluid) of animals is called [[serology]].<ref name="pmid32342927">{{cite journal |vauthors=Zainol Rashid Z, Othman SN, Abdul Samat MN, Ali UK, Wong KK |title=Diagnostic performance of COVID-19 serology assays |journal=The Malaysian Journal of Pathology |volume=42 |issue=1 |pages=13–21 |date=April 2020 |pmid=32342927 |doi= |url=}}</ref> Once an antibody–reaction has taken place in a test, other methods are needed to confirm this. Older methods included [[complement fixation test]]s,<ref name="pmid1365549">{{cite journal |vauthors=Swack NS, Gahan TF, Hausler WJ |title=The present status of the complement fixation test in viral serodiagnosis |journal=Infectious Agents and Disease |volume=1 |issue=4 |pages=219–24 |date=August 1992 |pmid=1365549 |doi= |url=}}</ref> [[Hemagglutination assay|hemagglutination inhibition]] and [[Neutralizing antibody|virus neutralisation]].<ref name="pmid21887208">{{cite journal |vauthors=Smith TJ |title=Structural studies on antibody recognition and neutralization of viruses |journal=Current Opinion in Virology |volume=1 |issue=2 |pages=150–6 |date=August 2011 |pmid=21887208 |pmc=3163491 |doi=10.1016/j.coviro.2011.05.020 |url=}}</ref> Newer methods use [[ELISA|enzyme immunoassay]]s (EIA).<ref name="pmid22185616">{{cite journal |vauthors=Mahony JB, Petrich A, Smieja M |title=Molecular diagnosis of respiratory virus infections |journal=Critical Reviews in Clinical Laboratory Sciences |volume=48 |issue=5–6 |pages=217–49 |date=2011 |pmid=22185616 |doi=10.3109/10408363.2011.640976 |s2cid=24960083 |url=}}</ref>

In the years before [[Polymerase chain reaction|PCR]] was invented [[immunofluorescence]] was used to quickly confirm viral infections. It is an infectivity assay that is virus species specific because antibodies are used. The antibodies are tagged with a dye that is luminescencent and when using an optical microscope with a modified light source, infected cells glow in the dark.<ref name="pmid32197545">{{cite journal |vauthors=AbuSalah MA, Gan SH, Al-Hatamleh MA, Irekeola AA, Shueb RH, Yean Yean C |title=Recent Advances in Diagnostic Approaches for Epstein-Barr Virus |journal=Pathogens |volume=9 |issue=3 |date=March 2020 |page=226 |pmid=32197545 |pmc=7157745 |doi=10.3390/pathogens9030226 |url=|doi-access=free }}</ref>

===Polymerase chain reaction (PCR) and other nucleic acid detection methods===
PCR is a mainstay method for detecting viruses in all species including plants and animals. It works by detecting traces of virus specific RNA or DNA. It is very sensitive and specific, but can be easily compromised by contamination. Most of the tests used in veterinary virology and medical virology are based on PCR or similar methods such as [[transcription mediated amplification]]. When a novel virus emerges, such as the covid coronavirus, a specific test can be devised quickly so long as the viral genome has been sequenced and unique regions of the viral DNA or RNA identified.<ref name="pmid32815744">{{cite journal |vauthors=Zhu H, Zhang H, Xu Y, Laššáková S, Korabečná M, Neužil P |title=PCR past, present and future |journal=BioTechniques |volume=69 |issue=4 |pages=317–325 |date=October 2020 |pmid=32815744 |pmc=7439763 |doi=10.2144/btn-2020-0057 |url=}}</ref> The invention of [[microfluidic]] tests as allowed for most of these tests to be automated,<ref name="pmid35300739">{{cite journal |vauthors=Wang X, Hong XZ, Li YW, Li Y, Wang J, Chen P, Liu BF |title=Microfluidics-based strategies for molecular diagnostics of infectious diseases |journal=Military Medical Research |volume=9 |issue=1 |pages=11 |date=March 2022 |pmid=35300739 |pmc=8930194 |doi=10.1186/s40779-022-00374-3 |url= |doi-access=free }}</ref> Despite its specificity and sensitivity, PCR has a disadvantage in that it does not differentiate infectious and non-infectious viruses and "tests of cure" have to be delayed for up to 21 days to allow for residual viral nucleic acid to clear from the site of the infection.<ref name="pmid33219449">{{cite journal |vauthors=Benzigar MR, Bhattacharjee R, Baharfar M, Liu G |title=Current methods for diagnosis of human coronaviruses: pros and cons |journal=Analytical and Bioanalytical Chemistry |volume=413 |issue=9 |pages=2311–2330 |date=April 2021 |pmid=33219449 |pmc=7679240 |doi=10.1007/s00216-020-03046-0 |url=}}</ref>

===Diagnostic tests===
In laboratories many of the diagnostic test for detecting viruses are nucleic acid amplification methods such as PCR. Some tests detect the viruses or their components as these include electron microscopy and [[ELISA|enzyme-immunoassays]]. The so-called "home" or "self"-testing gadgets are usually [[COVID-19 testing#Antigen tests|lateral flow tests]], which detect the virus using a tagged [[monoclonal antibody]].<ref>{{Citation|last1=Burrell|first1=Christopher J.|title=Chapter 10 - Laboratory Diagnosis of Virus Diseases|date=2017-01-01|work=Fenner and White's Medical Virology (Fifth Edition)|pages=135–154|editor-last=Burrell|editor-first=Christopher J.|place=London|publisher=Academic Press|language=en|doi=10.1016/b978-0-12-375156-0.00010-2|isbn=978-0-12-375156-0|pmc=7149825|last2=Howard|first2=Colin R.|last3=Murphy|first3=Frederick A.|editor2-last=Howard|editor2-first=Colin R.|editor3-last=Murphy|editor3-first=Frederick A.}}</ref> These are also used in agriculture, food and environmental sciences.<ref name="pmid27365041">{{cite journal |vauthors=Koczula KM, Gallotta A |title=Lateral flow assays |journal=Essays in Biochemistry |volume=60 |issue=1 |pages=111–20 |date=June 2016 |pmid=27365041 |pmc=4986465 |doi=10.1042/EBC20150012 |url=}}</ref>