Daily Archives: May 15, 2020

The first reported case of COVID-19 was 13 March 2020 in Kenya and 10 weeks later, not a single genome is available publicly from any of the East African Community countries (Kenya, Tanzania, Burundi, Uganda, Rwanda, South Sudan). Why is it so? And why does it matter? Globally the main focus during this outbreak has been rapid COVID testing and not whole-genome sequencing. The team at Nextstrain has highlighted the utility of whole-genome sequencing in addition to rapid testing. We have presented below some of the challenges to obtaining whole genomes in East Africa and most importantly we have suggested a way forward.

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As a diagnostic, whole genomes are critical. Sequences confirm the identity of the disease-causing pathogen and can be further used for studying diversity, tracing movement of virus strains, designing models that can predict the disease spread and to better understand the enemy. A recent French study in bioRxiv has claimed the SARS-CoV-2 strain in France was not imported from China. This highlights the importance of a sequencing initiative to be able to properly trace the progress of the pandemic in every setting the Icelandic approach.

Real-time data are very important because they serve as a diagnostic test that guides quick patient management and decision-making from an epidemiological standpoint; and genomics would provide further tools in designing therapeutic approaches.

Over the years, millions of USD have been spent building genomic sequencing facilities in East Africa. In Kenya, Biosciences for east and central Africa (plant and animal) and KEMRIWellcome Trust (both Nairobi, Kenya) (human health) are partnerships with national governments and international funders but to date neither have delivered a genome.

In Uganda, the Uganda Virus Research Institute (UVRI; Entebbe, Uganda), is a centre of excellence in virus research with the human and infrastructural capacity and international support for genome sequencing. However, UVRI has also not yet delivered a single SARS-CoV-2 genome.

Tanzania has a different landscape. There are no large international sequencing facilities, but the national research organizations, universities and hospitals like Muhimbili National Hospital (Dar es Salaam, Tanzania) and the Sokoine University of Agriculture (SUA; Morogoro, Tanzania) have various platforms such as the Illumina (CA, USA) MiSeq, HiSeq and the Oxford Nanopore MinION. They too have not yet generated any SARS-CoV-2 genomes.

So why have none of these institutions with the sequencing infrastructure and support in Kenya, Tanzania and Uganda not delivered the much-needed SARS-CoV-2 genomes yet?

Taking the highest tech genomics tools to the farmers in East Africa

DNA sequencing in Africa is currently a laborious task requiring researchers to send data to a centralized sequencing lab in Kenya or to await results from overseas. Here, Laura Boykin tells her story of working with the Tanzanian Agricultural Research Institute.

For Kenya, the biggest hurdle is a lack of partnerships. So far, all the work on COVID-19 is handled solely by the Ministry of Health (MoH; Nairobi, Kenya). Accordingly, there has been no access to samples considering also that this disease is highly infectious and these samples need to be handled in biosafety level 4 labs. Due to poor partnerships (aka poor coordination), the work is largely being done in KEMRI and private medical labs such as Lancet. The other limitations are:

Power, computers, internet and PCR machines are not a challenge.

The sequencing capacity is there especially in research and academic institutes; the SUA has the Thermo Fisher Scientific (MA, USA) Ion Torrent that they use for foot and mouth disease and other animal research, the Kilimanjaro Clinical research Institute (KCRI, Moshi, Tanzania) has the Illumina MiSEQ (I have seen this personally) which they use for their tuberculosis research; the Government Chemist Laboratory Authority (Dar es Salaam, Tanzania) has a genetic analyzer and was able to acquire the Illumina HiSEQ, which they use for their forensic studies; the National Health Laboratory (Dar es Salaam, Tanzania) also has a genetic analyzer. There are two laboratories which are capable of sequencing using Oxford Nanopore Technologies (Oxford, UK). These are Muhimbili national hospital and the SUA in collaboration with the NHL. There were no funds to do the sequencing at the beginning of the outbreak but now the SUA has secured some funds to sequence, Muhimbili might get a donation to do so too. Another laboratory that is capable of sequencing but does not have the funds to do so is the KCRI. Capacity and skills are not a problem. However, in a government setting and in most institutes, employees are given specific tasks as per institute mandate. Its true that we have many people trained in sequencing, but some are outside government settings/employment and some of those who are in government employment are not in clinical research. For example, the cassava disease diagnostic team was focused on agricultural research. Some of the trained people are not trained to handle clinical samples. So clearly there is a disconnect between clinical and agricultural disease diagnostic techniques.

Another challenge is lack of local partnerships (internal collaborations among different institutes in Tanzania). There are no good networks that connect healthcare facilities with research and academic institutes. Most healthcare facilities do not have the critical mass of trained experts in sequencing and due to their mandates and the sheer heaviness of their routine workload, they rarely have the bandwidth to pursue research regularly. Herein comes the need to forge strong links between the two that would have been in prime position to address this pandemic. Unfortunately it has not been easy; from my personal experience there are a lot of territorial issues at play that are hard to overcome. Perhaps this pandemic might bring a change in mindset.

Another challenge is global but is felt more in countries like Tanzania; inadequate funding for R&D. While the government, through the Tanzania Commission for Science and Technology (COSTECH) and other institutes, provides for R&D funding, it is still limited especially when compared to the costs of running genomics experiments. External funding is always difficult especially for researchers who are not part of a consortium led by PIs from Europe and/or North America. This has helped establish centers but has meant that the moment funding runs out the lab is less active, the reagents and consumables run out and equipment ends up in disuse.

There appears to be a lack of awareness among policy makers and/or not enough initiative from the local scientists working in this field to inform our policy makers about the importance of whole genome sequencing for management of COVID-19. Since most sequencing initiatives in the country are led by foreign consortia (which we feel needs to change) led from either Europe or North America it is possible that the benefits from such projects are rarely seen by policy makers in Tanzania. We see there needs to be a clear link between the governments and local scientists to work on the same matters from different perspectives. We hope the donated research reagents to the African CDC will reach the institutes as soon as they arrive the airport without customs delays.

There is both human and infrastructural capacity in sequencing at UVRI and the Medical Research Council all based at Entebbe, Uganda. However, the COVID-19 genomes are not yet out in the public arena.

There are computers, access to internet, power and the supplies required to carry out PCR testing and analysis of coronavirus/COVID-19 infections, which were initially provided by the UVRI through its running projects and currently with the support of the government. However, more supplies would be needed to monitor the entry and spread of the virus in the communities.

As of today, it is the sole responsibility of the Ministry of Health (Kampala, Uganda) as the mandated institution of government to lead all COVID-19 pandemic-related issues. This includes checking for possible cases with suspected symptoms, isolation/quarantine, collecting samples, sample analysis and announcement of outcomes of testing and treatment. In addition, task forces were established to coordinate COVID-19-related issues at national, regional and district level. The laboratory analysis of the suspected COVID-19 samples is carried out by UVRI. Although there are other institutions with both human and infrastructural capacity in molecular biology and disease diagnostics, there are limited partnerships on widening the testing for COVID-19 in the country to involve the private sector. This may be partly due to the highly infectious nature of the disease and the requirement to carry out the laboratory testing and analysis in a biosafety level 4 containment facility such as UVRI. However, there are some partnerships within the private sector in management of the disease.

Insight into SARS-CoV-2 genome spells good news for vaccine development

Infectious disease researchers have identified just five SARS-CoV-2 gene variants, suggesting a vaccine for COVID-19 could be highly effective.

This article is written by East African Scientists and international partners who have been working for years on collaborative research projects, including The Cassava Virus Action Project, around managing emerging plant virus disease pandemics using novel molecular diagnostics and genomics. The team was disheartened to watch COVID-19 arrive and spread in East African countries, where they have successfully partnered to build capacity in rapid plant virus diagnostics and genome sequencing using novel portable technologies such as the Oxford Nanopore MinION, which have not been put to good use in the fight against the pandemic.

Professor Elijah Ateka Molecular Biologist

Dr. Joseph Ndunguru Molecular Plant Pathologist

Dr. Daniel Maeda Molecular and Cellular Biologist (Health Focus)

Mr. Charles Kayuki Molecular Biologist

Dr. Peter Sseruwagi Molecular disease epidemiologist

Dr. Laura M. Boykin Computational Biologist

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Where Are The SARS-CoV-2 Genomes From East Africa? - BioTechniques.com

Viruses may seem like cunning villains, purposefully mutating to increasingly deadlier forms to outwit their human hosts. In reality, a lot of what happens with a virus is completely random. This randomness can make figuring out where a virus came from, how it spreads and what makes it tick especially tricky. For SARS-CoV-2, the new virus that causes COVID-19, scientists are looking to its genome for answers to some of these questions.

Researchers were recently able to determine that New York City may have been the original epicenter of the U.S. epidemic and that those initial cases were likely imported from Europe. They can tell this by looking at the genome and its sequence and seeing how they are similar or different, explains Adam Lauring, M.D., Ph.D., associate professor of microbiology and immunology and infectious disease.

Armed with virus samples taken from people with COVID-19, virologists and epidemiologists create what is known as a phylogenetic tree. This viral family tree lines up the genetic codes from each sample of virus to see whos related to whom.

Based on the genetic sequences and time of collection, you can start to paint a picture of how the virus moves through a population. The earliest [virus samples] in New York were more similar to the ones from people in Europe who were infected. You start with the dates, then look at the sequences and figure thats the most likely scenario, says Lauring. Its not foolproof, though. Theres always uncertainty.

A real world example of this uncertainty came to light with a study posted online in April, which described the deaths of two people from COVID-19 in Santa Clara, California weeks earlier than the virus was thought to be in California. What this tells us is that theres definitely missing data, says Lauring. This begs the question, he says, of where did those cases came from and how long the virus was spreading before the outbreak was recognized.

SEE ALSO: Seeking Medical Care During COVID-19

Researchers are also looking at the SARS-CoV-2 genome for clues about its true origin: the animal that infected the first person. So far, bats appear to be the most likely suspect. Looking at the phylogenetic tree, we see that a bat coronavirus is the closest relative to SARS-CoV-2, sharing around 96% of their genomes, says Lauring. But that too, is not the full story. Another animal, a small, scaly-skinned mammal called a pangolin, has been implicated as well.

The spike protein in SARS-CoV-2, the main protein on the surface that binds to the cells receptor and how the virus gets into the cell, is similar to a pangolin coronavirus spike protein, says Lauring. Its almost like, when you tell a person he has his fathers nose. That feature is similar, but across features the father and child may not look very similar. Coronaviruses, like a lot of other viruses, swap genes around.

These swaps are examples of mutations, which are common in RNA viruses like SARS-CoV-2. Laurings lab focuses on mutations in influenza, the RNA virus behind the infamous 1918 Spanish flu pandemic. Understanding how influenza mutates is critical for making decisions about the annual influenza vaccine. RNA viruses mutate relatively quickly because they lack a proofreading mechanism to look for and repair errors during replication.

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However, SARS-CoV-2 and its coronavirus cousins are unique among RNA viruses, because they have a proofreading enzyme. The coronavirus genomes are three times longer than youd expect them to be, and the presence of the proofreading enzyme explains that nicely, says Katherine Spindler, Ph.D., professor in the department of microbiology and immunology. Spindler is a host for the podcast This Week in Virology, which examines the latest science around SARS-CoV-2 and other viruses.

With this enzyme, the virus can make a few more errors and not have it be lethal for the virus. As a result, SARS-CoV-2 mutates more slowly than other RNA viruses. Spindler notes that only about 20 mutations have been retained in the genome so far since the beginning of 2020, despite the billions of times the virus has replicated.

SEE ALSO: Keeping Our Patients Safe During COVID-19

Even with its relatively slow mutation rate, mutations present in each persons SARS-CoV-2 genome allows researchers to do genetic tracing in real time, says Lauring. His lab hopes to study the virus genome more closely to look at how the virus is transmitted in healthcare settings and communities.

He stresses that just because a virus mutates doesnt mean the mutations are making it stronger, more likely to be transmitted, or that it will be tougher to develop a vaccine. My hunch is evolution wont be the biggest challenge in developing a vaccine. There are viruses that evolve relatively quickly for which we do have vaccines, for example polio, measles, mumps, Ebola, hepatitis A, notes Lauring.

Spindler adds that the fact that were seeing a variety of COVID-19 symptoms doesnt mean there are different mutant strains. Every new symptom that comes along, from COVID toes and skin rashes to blood clots, are likely just additional manifestations of the virus as it infects so many different people, she says. Figuring out the mysteries of SARS-CoV-2 will take years of experimental work, she says.

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What the SARS-CoV-2 Genome Reveals - Michigan Medicine

The discovery of a huge number of long non-protein coding RNAs, aka lncRNAs, inthe mammalian genome was a major surprise of the recent large-scale genomics projects. Aninternational team including a bioinformatician from the Research Center of Biotechnology of the Russian Academy of Sciences, and the Moscow Institute of Physics and Technology has developed areliablemethodfor assessing therole of such RNAs. Thenew technique and the data obtained with it allow generating important hypotheses on how chromatin is composed and regulated, aswell as identifying the specific functions of lncRNAs.

Presented inNature Communications, the technology is called RADICL-seq and enables comprehensive mapping of each RNA, captured while interacting with all thegenomic regions that it targets, where many RNAs are likely to be important forgenome regulation and structure maintenance.RNA and gene regulationIt was previously believed that RNA functions mostly as an intermediary in building proteins based on a DNA template, with very rare exceptions such as ribosomal RNAs. However, with the development of genomic analysis, it turned out that not all DNA regions encode RNA, and not all transcribed RNA encodes proteins.

Although the number of noncoding RNAs and those that encode proteins is about the same, the function of most noncoding RNA is still not entirely clear.

Every type of cell has its own set of active genes, resulting in the production ofspecific proteins. This makes a brain cell different from a blood cell of the same organism despite both sharing the same DNA. Scientists are now coming to theconclusion that RNA is one of the factors that determine which genes are expressed, or active.

Long noncoding RNAs are known to interact with chromatin DNA tightly packaged with proteins. Chromatin has the ability to change its conformation, or shape, so that certain genes are either exposed for transcription or concealed. Long noncoding RNAs contribute to this conformation change and the resulting change in gene activity by interacting with certain chromatin regions. To understand the regulatory potential of RNA in addition to it being a template for protein synthesis it is important to know which chromatin region any given RNA interacts with.

How it works

RNAs interact with chromatin inside the cell nucleus by binding tochromatin-associated proteins that fold a DNA molecule. There are several technologies that can map such RNA-chromatin interactions. However, all of them have significant limitations. They tend to miss interactions, or require a lot of input material, or disrupt the nuclear structure.

Toaddress these shortcomings, a RIKEN-led team has presented a new method: RNA and DNA Interacting Complexes Ligated and Sequenced, or RADICL-seq for short. The technique produces more accurate results and keeps the cells intact upuntil theRNA-chromatin contacts are ligated.

The main idea of the RADICL-seq method is the following. First, the RNA is crosslinked to proteins located close to it in the nucleus of cells with formaldehyde. Then, DNA is cut into pieces by digesting it with a special protein. After that, thetechnology employs RNaseH treatment to reduce ribosomal RNA content, thus increasing the accuracy of the result. Then, by using a bridge adapter (amolecule with single-stranded and double-stranded ends) the proximal DNA and RNA are ligated. After the reversal of crosslinks, the RNA-adapter-DNA chimera is converted to double-stranded DNA for sequencing, revealing the sequence of the ligated RNA and DNA.

Decoding the noncoding

Incomparison with other existing methods, RADICL-seq mapped RNA-chromatin interactions with a higher accuracy. Moreover, the superior resolution ofthetechnology allowed the team to detect chromatin interactions not only with thenoncoding but also with the coding RNAs, including those found far from their transcription locus. The research confirmed that long noncoding RNAs play animportant role in the regulation of gene expression occurring at a considerable distance from the regulated gene.

This technology can also be used to study cell type-specific RNA-chromatin interactions. The scientists proved it by looking at two noncoding RNAs in a mouse cell, one of them possibly associated with schizophrenia. They found that aninteraction pattern between chromatin and those RNAs in two different cells theembryonic stem cell and the oligodendrocyte progenitor cell correlated with preferential gene expression in those cell types.

The new methods flexibility means scientists can gather additional biological information by modifying the experiment. In particular, this technology can make it possible to identify direct RNA-DNA interactions not mediated by chromatin proteins. The analysis performed by bioinformaticians from the Research Center ofBiotechnology and MIPT showed that not only the standard double helix interactions between DNA and RNA but also those involving RNA-DNA triplexes could participate in gene regulation. Also, such interactions highlight the significance of noncoding RNA in protein targeting to particular gene loci.

We are planning to conduct further research on the role of RNA in the regulation ofgene expression, chromatin remodeling, and ultimately, cell identity. Hopefully, we will be able to regulate genes by using these noncoding RNAs in the near future. This can be especially helpful for treating diseases, saysYulia Medvedeva, who leads the Regulatory Transcriptomics and Epigenomics group at the Research Center of Biotechnology, RAS, and heads the Lab of Bioinformatics for Cell Technologies at MIPT. She also manages the grant project supported by the Russian Science Foundation, which co-funded the study.

Reference:Bonetti, A., Agostini, F., Suzuki, A.M. et al. RADICL-seq identifies general and cell typespecific principles of genome-wide RNA-chromatin interactions. Nat Commun 11, 1018 (2020). https://doi.org/10.1038/s41467-020-14337-6.

This article has been republished from the following materials. Note: material may have been edited for length and content. For further information, please contact the cited source.

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A Method for Assessing the Role of Long Non-protein Coding RNAs - Technology Networks

The 15th Annual Endangered Species Day sees more than 31,000 species around the world threatened with extinction, according to the IUCN Red List. That is almost one third of the groups of animals listed on their website.

Researchers are now using a vast remote database to help protect endangered species. This genomic library allows the team to access vital datasets more efficiently than ever before, thanks to a collaboration between the University of Sydney and Amazon Web Services (AWS).

Were often working with more than a billion pieces of jigsaw puzzle and no guide, says senior research manager Dr Carolyn Hogg. The software now helps condense the scientists work enormously, enabling them to analyse massive amounts of data in minutes rather than hours - regardless of where they are in the world.

In the long term, the researchers aim to share this genome data publicly. The ultimate goal would be to create a universal genomic library and tools that other researchers and conservation managers can access in order to make science-based decisions, adds Dr Hogg.

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Endangered species could be saved with this tech-based solution - Euronews