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Discussion: Chinese Academy of Sciences (CAS) in Wuhan has been working with bats and coronavirus for many years - DNA manipulations, cloning....

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  • #16
    Also please see:

    nCoV-2019 Spike Protein Receptor Binding Domain Shares High Amino Acid Identity With a Coronavirus Recovered from a Pangolin Viral Metagenomic Dataset

    jan 30, 2020

    An outbreak of respiratory illness caused by a novel coronavirus (nCoV-2019, NC_045512.2 52) first identified in Wuhan China has resulted in over seven thousand confirmed cases.


    https://flutrackers.com/forum/forum/...emains-unclear


    Wondering if people can transmit SARS-like coronaviruses to pangolins? Supposedly numerous wild life was traded at the Wuhan Seafood market. Is pangolin the reservoir for COVID-19 or are humans?

    I am not a virologist but it seems that if humans can transmit disease to dogs then why not other mammals? We do not know how long COVID-19 was "in the making" in China. It may have been months - circulating inefficiently - stops and starts - until mutated through enough generations to become efficient human-to-human.

    Looking forward to the tribunal on this.


    Comment


    • Kathy
      Kathy commented
      Editing a comment
      Another interesting article on CoVs in pangolin:

      Liu, P.; Chen, W.; Chen, J.-P. Viral Metagenomics Revealed Sendai Virus and Coronavirus Infection of Malayan Pangolins (Manis javanica). Viruses 2019, 11, 979

      "All these Malayan pangolins were rescued by the Guangdong Wildlife Rescue Center, however, 16 died after extensive rescue efforts. Most of the dead pangolins had a swollen lung which contained a frothy liquid, as well as the symptom of pulmonary fibrosis, and in the minority of the dead ones, we observed hepatomegaly and splenomegaly."

      Most of the pangolins were also infected with Herpes virus, that, please correct me if I am wrong, present a furin cleavage site.

      I could not find CoVs sequences submitted in NCBI from this work, but I came to MT084071 (I already posted some information on this sequence here https://flutrackers.com/forum/forum/...genetics/page3), that is not properly sequenced and is most probably related to this work (see author list in the NCBI form below)

      LOCUS MT084071 27213 bp RNA linear VRL 20-FEB-2020 DEFINITION Pangolin coronavirus isolate MP789 genomic sequence. ACCESSION MT084071 VERSION MT084071.1 KEYWORDS . SOURCE Pangolin coronavirus ORGANISM Pangolin coronavirus Viruses; Riboviria; Nidovirales; Cornidovirineae; Coronaviridae; Orthocoronavirinae; Betacoronavirus; unclassified Betacoronavirus. REFERENCE 1 (bases 1 to 27213) AUTHORS Jiang,J.-Z., Liu,P. and Chen,J.-P. TITLE Are pangolins an intermediate host of the 2019 novel coronavirus (2019-nCoV) ? JOURNAL Unpublished REFERENCE 2 (bases 1 to 27213) AUTHORS Jiang,J.-Z., Liu,P. and Chen,J.-P. TITLE Direct Submission JOURNAL Submitted (13-FEB-2020) SCSFRI, South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences (SCSFRI, CAFS), 231# Xingangxi Road, Haizhu District, Guangzhou, Guangdong 510300, China.

      Also this sequence was submitted after the outbreak of SARS-CoV2. I do not know how the Chinese Academy of Fishery Sciences could be interested in the topic.

      More details on MT084071 are given in: Genetic evolution analysis of 2019 novel coronavirus and coronavirus from other species Chun Li, Yanling Yang, Linzhu Ren
      https://www.sciencedirect.com/scienc...167?via%3Dihub

      They mention: "Moreover, SARS-CoV-2 and pangolin coronavirus is also highly related (Lam et al., 2020; Xiao et al., 2020; Zhang et al., 2020b), but due to incomplete sequence of pangolin coronavirus published in GenBank, reasonable analysis cannot be carried out in this study".

  • #17
    I had a quick look at Pangolins a few weeks ago as an intermediary host. They are genetically on a branch with cats and dogs and they have ACE2 receptors which are a close match, but ACE2 is a common to many species. However they are critically endangered with very small population size, they are secretive by nature and like a wooded hill side habitat. The main population is in the south and well away from the epicenter. While possible this makes it poor potential host, something more common, even if seldom infected, would seem more likely.
    Last edited by JJackson; February 16th, 2020, 07:39 PM.

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    • #18
      hat tip Shiloh


      Apparently pulled from researchgate but preserved by Wayback Machine.

      https://web.archive.org/web/20200214...oV_coronavirus





      Click image for larger version

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      Comment


      • #19
        Source: https://www.voachinese.com/a/5290767.html

        Hong Kong Fengyun (February 16, 2020) Wuhan P4 laboratory confirmed "poison"? Xi Jinping pushes "Biosecurity Law"
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        Hong Kong Fengyun (February 16, 2020) Wuhan P4 laboratory confirmed "poison"? Xi Jinping pushes "Biosecurity Law"
        share it


        Washington —

        As Wuhan's pneumonia epidemic raged, Xi Jinping made a special mention of accelerating the promotion of biosafety legislation and incorporating biosafety into the national security system when he addressed a meeting of the Central Committee of Comprehensive and Deepening Reform on February 14, attracting the attention of all circles. New crown virus may come from the laboratory? Why did Xi Jiping push for the biosafety law at this time? Hong Kong Fengyun invited Su Yiren, former director of the Taiwan Bureau of Disease Management and lecturer of the Department of Technology of Nantai University of Science and Technology to analyze.

        P4 laboratory needs careful management

        Su Yiren: The P4 laboratory is a very important facility in the past in the production of vaccines or animal transmission models, or even in bioterrorism. So every country in this place is very careful to do management or legislation. According to the materials published by the professors of South China University of Technology, I am also very surprised. They have done a lot of bat experiments. The CDC, the Virus Research Institute, or the P4 laboratory in Wuhan are doing advanced work in several places. Experiment, if it is useful to the P4 laboratory, this place is the point we have been concerned about. In particular, some researchers were sprayed with blood or urine from bats. This part is actually the most important point for the P4 laboratory when doing some new viruses. It is the most important in the human protection or software supporting facilities of the P4 laboratory. of.

        Wuhan P4 laboratory's safety establishment does not meet regulations

        Su Yiren: The establishment of a P4 laboratory requires very high standards and requires national level management. It's not just hardware equipment. Generally speaking, there is a certain standard for the living environment. How many kilometers away can't be close to the people. Therefore, the distance between the South China Seafood Market and Wuhan P4 Lab is 280 meters. I was surprised when I heard this. I should not set up a P4 Lab at this distance. I don't know the background of the original establishment, but this is not in accordance with the regulations.

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        • #20
          all the genbank-known sequences are still very distant from covid , including RaTG13 .
          No signs for a lab-escape here

          They might have found covid in pangolins or bats and kept it secret, worked with it,
          maybe even manipulated the cleavage site and then it escaped.
          But that's not typical for their research, which requires publishing.

          They may have found the virus (in Yunnan ?), did some first testing, immediately recognised
          its military potential and kept everything secret while growing it, testing it further
          I'm interested in expert panflu damage estimates
          my current links: [url]http://bit.ly/hFI7H[/url] ILI-charts: [url]http://bit.ly/CcRgT[/url]

          Comment


          • #21
            Originally posted by sharon sanders View Post
            hat tip Shiloh


            Apparently pulled from researchgate but preserved by Wayback Machine.

            https://web.archive.org/web/20200214...oV_coronavirus





            Click image for larger version

Name:	originsofcov.PNG
Views:	744
Size:	120.1 KB
ID:	829777

            Full paper here:

            Preprint Paper: The possible origins of 2019-nCov coronavirus

            Comment


            • #22
              Originally posted by gsgs View Post
              all the genbank-known sequences are still very distant from covid , including RaTG13 .
              No signs for a lab-escape here

              They might have found covid in pangolins or bats and kept it secret, worked with it,
              maybe even manipulated the cleavage site and then it escaped.
              But that's not typical for their research, which requires publishing.

              They may have found the virus (in Yunnan ?), did some first testing, immediately recognised
              its military potential and kept everything secret while growing it, testing it further
              What possible military potential does this virus offer? It is highly infectious, but slow acting and only moderately lethal. I struggle to envision a practical military use for it.

              It seems more likely that this was just another research effort to try to understand these SARS related bat viruses. We know the scientists have at times been very blase about their study subjects, even carrying deadly germs in their airplane luggage, so the possibility of a slip up at a facility is quite real.
              Separately, I'd thought the virus was well over 90% identical to an earlier horseshoe bat carried specimen, but with some new sequences included which make it so dangerous. There does not yet appear to be a consensus as to where those new sequences come from.

              Comment


              • #23
                to demonstrate to the world the superiority of Chinese communism with strong government over Western democracies
                with freedom and reluctant public health measures.

                Xi : China will be stronger than before after this (or similar)
                I'm interested in expert panflu damage estimates
                my current links: [url]http://bit.ly/hFI7H[/url] ILI-charts: [url]http://bit.ly/CcRgT[/url]

                Comment


                • #24
                  they had 96% similarity with RatG13 from 2013. Not enough to assume a lab-escape, even if the escape already happened in 2013.
                  You can't apply these changes in a lab so to hide things - someone will discover a natural virus and the deceiving becomes obvious.

                  I assume that internationally the search for similar viruses in animals is being intensified now.
                  When you hide a virus , others may find a similar one ...

                  In old days they replicated the viruses through many generations in the lab and mutations were added.
                  Nowadays they freeze them in liquid nitrogen to preserve them (afaik, what I'd read, I'm not a lab-person)
                  I'm interested in expert panflu damage estimates
                  my current links: [url]http://bit.ly/hFI7H[/url] ILI-charts: [url]http://bit.ly/CcRgT[/url]

                  Comment


                  • #25
                    Translation Google

                    Pneumonia outbreak: ambiguous "Patient Zero" and virus source dispute

                    Wang Yiqing
                    BBC Chinese
                    February 18, 2020

                    China is working to curb the spread of the new coronavirus pneumonia, but where exactly the outbreak has started is still foggy. A doctor who was involved in treating the first new crown patient currently known to the public told the BBC that the patient was a patient with cerebral infarction in his 70s at home.

                    This is the first time that information about the early patient has been made public. He is believed to have developed the disease on December 1st, almost a week earlier than the patient previously reported by Wuhan officials on December 8th. It is worth noting that he has no history of exposure to the South China seafood market.

                    The wildlife trade in the South China Seafood Market in Wuhan has been suspected to have been the culprit for the epidemic, but the experience of the old man is challenging that conclusion. Scholars and netizens have questioned whether the virus has other potential sources, including whether it may be related to Wuhan Virus Research Institute.

                    Earliest known patient
                    The new outbreak of coronavirus pneumonia in Wuhan, which has infected more than 70,000 people and killed nearly 2,000 people in China, remains unclear as to how the virus initially crossed the species barrier and transmitted from animals to people. The key to understanding the virus's path of transmission is the identity of the first patient and how he became infected.

                    The Wuhan Municipal Health and Health Commission stated in a notification that the onset of the first case of new coronary pneumonia was December 8 last year, but an international authoritative medical journal "The Lancet" published on January 24 by A thesis written by Huang Chaolin and other deputy directors of Wuhan Jinyintan Hospital, who admitted to treating patients with new coronary pneumonia, pushed the first patient's onset time to December 1. The paper was written by nearly 30 researchers in Chinese medical institutions, and a considerable number of them are working in the field of treating patients with new crowns.

                    Dr. Wenjuan Wu, director of the Intensive Care Unit (ICU) at Jinyintan Hospital and one of the authors of the above papers, told the BBC on Monday (February 17) that the patient who developed the disease on December 1 was a man over seventies . The onset time on December 1 was a conclusion drawn from the epidemiological survey of the family members.

                    "This patient has a cerebral infarction and senile dementia, and the condition was very bad when he was sent over," Wu Wenjuan said. She declined to disclose the patient's last name.

                    It is reported that after the onset of the disease, the patient was first admitted to another hospital in Wuhan, but as the condition worsened, he was transferred to Jinyintan Hospital on December 29. At the time, Huang Chaolin and Wu Wenjuan were on the scene.

                    Wu Wenjuan said the old man had been sick at home before and had not visited the South China Seafood Market, a trading market in Wuhan that sells seafood and game products. Because a large number of merchants in this market became ill at the beginning of the outbreak, it was once considered the origin of the outbreak.

                    "He lives far from four or five stops (bus stops) at the seafood market," Wu Wenjuan said. "And because he was sick, he basically didn't go out."

                    According to a paper published in The Lancet, the elderly's family members did not develop fever or respiratory symptoms after the onset of illness, and there was no epidemiological link between them and subsequent patients. Ten days after his onset, three other people developed symptoms, two of whom had no history of exposure to the South China seafood market.

                    The BBC has not been able to independently verify this information.

                    Source of contradiction

                    The BBC asked Wu Wenjuan, "Why is this patient who has been living at home for a long time and has never been to the South China Seafood Market infected with this newly discovered virus? Is there any other possible source of infection?"

                    She avoided the problem.

                    "What you are asking is exactly the direction of our next research," said Wu Wenjuan.

                    But it is obvious that this contradicts the previous widely speculated that the epidemic was a large number of operators or game buyers of the South China seafood market that transmitted the virus directly to the South China seafood market.

                    Huang Chaolin previously said in an interview with the Chinese media "Caixin.com": "From the perspective of the current disease situation, the seafood market is not the only source of exposure ... it is multi-source," but he believes that the virus has It is likely to still come from wild animals.

                    This wild animal is now generally considered a bat. As early as February 3, Nature, an international authoritative academic journal, Nature published a research paper on the origin of the new crown virus by Shi Zhengli, a researcher at the Wuhan Institute of Virology, Chinese Academy of Sciences, who believed that bats were a possible source of the pneumonia epidemic.

                    At a press conference on Saturday (February 15), officials from the Chinese Ministry of Science and Technology reiterated that bats are still the most likely source of the new crown virus, but pangolin may be one of the intermediate hosts of the new crown virus.

                    This reminds people of the SARS epidemic that prevailed in China from 2002 to 2003. In this outbreak, scientists first thought that the virus originated from civet, but eventually the disease was identified as a bat. But to this day, the first SARS patient-a chef in Heyuan, Guangdong-still insists he has not been exposed to these wild animals.

                    Wuhan Virus Institute in Controversy
                    Are there other possibilities?

                    The Wuhan Virus Research Institute mentioned above is a focus that has recently been questioned by public opinion. As Chinese officials have been extremely secretive about the identity of the first patient, netizens have been questioning whether the first patient was an employee of the Wuhan Virus Research Institute.

                    For example, last Saturday (February 15), the rumor that "Wu Yanling, a female graduate student at Wuhan Institute of Virology, is a patient with New Coronary Pneumonia No. 0" was widely circulated in Chinese social media. "Patient Zero" generally refers to the first patient who became infected with the virus and started to spread the virus.

                    The institute and Huang Yanling's company had a "rumor rumor" on Sunday, saying that the lady "has been working and living in other provinces since graduation." Shi Zhengli, a researcher at the institute, told the media, "None of us have been infected with the virus, and we have zero infection."

                    In addition, as early as the outbreak of the outbreak, a paper from the Indian Institute of Technology team questioned that the four independent inserts of the new crown virus "is unlikely to happen by accident in nature," which instantly made the outside world for "Contributing to the creation of biological and chemical weapons" conspiracy theory has been raging, but then the author of the article announced the withdrawal.

                    According to Rutgers University biologist Richard H. Ebright, according to the BBC, according to the current genome sequencing of the virus, there is no evidence that the virus has been artificially modified. In Nature, he expressed concern about a bat virus experiment at the Wuhan Institute of Virology.

                    But Eblatt added that this does not represent a possibility that the virus from the outbreak could enter the population due to a laboratory accident.

                    He said that genome sequencing showed that the outbreak virus was very close to the bat coronavirus RaTG13 collected in a cave in Yunnan in 2003 by the Wuhan Virus Research Institute, with a genome-wide homology of 96.2%.

                    "This means that the virus is currently known to exist in two places: a cave in Yunnan and a laboratory at the Wuhan Institute of Virology," Eblet said. "It has been stored at the Wuhan Institute of Virology since 2013. "

                    On February 14, Chinese President Xi Jinping proposed at a meeting that biosafety should be “incorporated into the national security system” and the introduction of a biosafety law be promoted as soon as possible. The next day, the Ministry of Science and Technology of China demanded “strengthening the management of laboratories, especially viruses”.

                    When asked if the first elderly person diagnosed had any relatives related to the Wuhan Virus Research Institute or the South China Seafood Market, Director Wu Wenjuan of Jinyintan Hospital said that "it is impossible to draw conclusions" now.

                    https://www.bbc.com/zhongwen/simp/chinese-news-51540821
                    "Safety and security don't just happen, they are the result of collective consensus and public investment. We owe our children, the most vulnerable citizens in our society, a life free of violence and fear."
                    -Nelson Mandela

                    Comment


                    • #27
                      Hattip Tetano

                      Virology Blog: Furin cleavage site in the SARS-CoV-2 coronavirus glycoprotein
                      Today, 12:31 PM
                      By Vincent Racaniello
                      ...
                      Examination of the protein sequence of the S glycoprotein of SARS-CoV-2 reveals the presence of a furin cleavage sequence (PRRARS|V). The CoV with the highest nucleotide sequence homology, isolated from a bat in Yunnan in 2013 (RaTG-13), does not have the furin cleavage sequence. Because furin proteases are abundant in the respiratory tract, it is possible that SARS-CoV-2 S glycoprotein is cleaved upon exit from epithelial cells and consequently can efficiently infect other cells. In contrast, the highly related bat CoV RaTG-13 does not have the furin cleavage site.

                      Whether or not the furin cleavage site within the S glycoprotein of SARS-CoV-2 is actually cleaved remains to be determined. Meanwhile, it is possible that the insertion of a furin cleavage site allowed a bat CoV to gain the ability to infect humans. The furin cleavage site might have been acquired by recombination with another virus possessing that site. This event could have happened thousands of years ago, or weeks ago. Upon introduction into a human – likely in an outdoor meat market – the virus began its epidemic spread.

                      Furins are also known to control infection by avian influenza A viruses, in which cleavage of the HA glycoprotein is needed for entry into the cell. Low-pathogenic avian influenza viruses contain a single basic amino acid at the cleavage site in the HA protein which is cleaved by proteases that are restricted to the respiratory tract. Insertion of a furin cleavage site in the HA of highly pathogenic avian H5N1 influenza viruses leads to replication in multiple tissues and higher pathogenicity, due to the distribution of furins in multiple tissues.

                      Acquisition of the furin cleavage site might be viewed as a ‘gain of function’ that enabled a bat CoV to jump into humans and begin its current epidemic spread.

                      http://www.virology.ws/2020/02/13/fu...-glycoprotein/

                      https://flutrackers.com/forum/forum/...s-glycoprotein
                      "Safety and security don't just happen, they are the result of collective consensus and public investment. We owe our children, the most vulnerable citizens in our society, a life free of violence and fear."
                      -Nelson Mandela

                      Comment


                      • #28
                        Translation Google

                        Scholar: New artificial pneumonia virus is highly likely

                        2020-02-22 14:43 Web News Agency / Taipei 22nd

                        Fang Qitai, a professor at the National Taiwan University School of Public Health, said today that academically , the new crown pneumonia virus may indeed be artificial, but administrative investigation is needed to reach a conclusion. In addition, if the virus is artificial, it means that there is no ecological place in nature, and the root may be broken in the future.

                        The Taiwan Society of Public Health held a "Newcastle Virus Public Health Community Epidemic Prevention Education Course" at the National Taiwan University today, inviting a number of public health experts to give lectures and broadcasting it on the Facebook fan page, attracting many teachers and students of public health related departments and people concerned about the epidemic Watch.

                        When Fang Qitai gave a lecture on "The Epidemiology of New Coronary Pneumonia," he quoted a number of viewpoints from overseas academia to discuss whether the new coronary pneumonia might be a man-made product of the laboratory and accidentally leaked out to cause disaster instead of natural evolution.

                        Fang Qitai said that many European and American scholars questioned that the outbreak of new crown pneumonia could not be separated from the New Crown Virus Research Institute of the Chinese Academy of Sciences. In its P4 laboratory, there are a large number of deadly viruses such as SARS and Ebola. Room culture has always been questioned.

                        Fang Qitai said that existing research literature pointed out that the new crown pneumonia virus and a bat virus RaTG13 retained by a new crown virus research institute have a similarity of 96%, causing academic discussion. However, from a virological point of view, 96% are not the same, and it may take more than 99% to count, but the key is the difference.

                        The French team compared the gene sequences and found that the new coronary pneumonia virus has 4 amino acids more than other coronaviruses, which may cause it to be more invasive, pathogenic, and transmissible than the original virus. Therefore, many people associate with it. Is it possible that some people think that SARS was too easy to be cracked 17 years ago, so they have developed an "upgraded version", and it can be done from the current technology.

                        Fang Qitai pointed out that most of the evolution in nature is a single point mutation, and it is unlikely that 4 amino acids will be added all at once. Therefore, academically speaking, new coronary pneumonia may indeed be artificially added in the laboratory; Sex is not without, but chances are very small.

                        Fang Qitai emphasized that at present, it can only be said that academically, the mutation of neo-pneumococcal pneumonia is unusual and may indeed be a man-made product. But in fact, internal administrative investigation is needed to find out laboratory records and other evidence. There are answers.

                        Fang Qitai believes that whether the new crown pneumonia is artificial will have an important impact on the future investigation and judgment of the epidemic. If it is an artificial virus, it means that it does not have its ecological position in nature, unlike influenza. Influenza is naturally evolved and is part of the ecology. Therefore, no one dares to say that it can block or eliminate the flu. Can reduce disasters and reduce severe deaths.

                        But there is no virus in nature, which means that when the last patient is cured, it can be cut off and disappear. The epidemic will eventually become a thing of the past, with the opportunity to disappear like SARS.

                        https://udn.com/news/story/6656/4363085
                        "Safety and security don't just happen, they are the result of collective consensus and public investment. We owe our children, the most vulnerable citizens in our society, a life free of violence and fear."
                        -Nelson Mandela

                        Comment


                        • #29
                          Originally posted by JJackson View Post
                          I had a quick look at Pangolins a few weeks ago as an intermediary host. They are genetically on a branch with cats and dogs and they have ACE2 receptors which are a close match, but ACE2 is a common to many species. However they are critically endangered with very small population size, they are secretive by nature and like a wooded hill side habitat. The main population is in the south and well away from the epicenter. While possible this makes it poor potential host, something more common, even if seldom infected, would seem more likely.
                          There's been rumors and this sounds like good confirmation that farming has been going on. Possibly the farming contributed but I think a human was the vector, maybe a farmer of bats or pangolins brought it to town. Even a bat researcher could have gotten infected by this in a cave.

                          https://www.theguardian.com/environm...-farm-industry
                          On a visit to Shaoguan, Guangdong province, last year, the Guardian and staff from CBCGDF saw a caged facility previously used for attempted breeding of the notoriously hard-to-breed pangolin.

                          While there were no longer pangolin at the site, several locals near the facility confirmed the species had been raised there, along with monkeys and other wildlife.
                          I did consider biotech. I don't care for all those proliferating animal labs. The best theory I could come up with based on that redacted paper about the 4 HIV sequence insertions, (gp120 and GAG?), was that this was an HIV vaccine candidate gone haywire. I found one paper from 2006 where coronaviruses were being researched as vectors to carry those HIV antigens. The research was from Europe but the primate lab in Wuhan is attracting teams from Europe because of the lax animal protection laws in China.
                          Still, there were no papers on the concept since then and they were considering using a mild human coronavirus. Even though common they said it would work because people don't develop protective immunity to the these viruses. But this novel virus seems too strange for anyone to have tried to use it as a vaccine vector. Why do that if a plain old cold virus will work?

                          So my opinion is that no sophisticated bioengineering was involved in creating this new human virus currently circulating.
                          “‘i love myself.’ the quietest. simplest. most powerful. revolution ever.” ---- nayyirah waheed

                          Avatar: Franz Marc, Liegender Hund im Schnee 1911 (My posts are not intended as advice or professional assessments of any kind.)

                          Comment


                          • #30
                            Emily
                            Genetic clock data gives the date of COVID human samples all coming from a single introduction a few months ago. The same calculations show divergence from either the closest bat or pangolin sequences as a couple of decades ago. No animal host has been found carrying the decades of lost viral evolution, but it must have been evolving somewhere. There is no evidence of any human interference in this virus' evolution and where close gene sequences (like the snakes) have been found it is due to common mechanisms, and genes, of ancient origin which can be found in most current life forms. The fact that so many drug trials are going on with existing drugs for diseases as diverse as flu, HIV and SARS is due to the same common mechanisms used for cell attachment, cleavage and fusion.

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