Abstract:
The Lassa Fever is a disease caused by the Lassa Virus, an old world arenavirus that is endemic in West Africa. In this paper, I will be examining Lassa Fever, its viral structure, symptoms, viral process, its transmission and treatment, relevant recent studies and reports, and its societal impact. The Lassa Fever is a zoonotic enveloped negative single stranded RNA virus, which fuses to and enters a cell. No vaccine currently exists, however, recently major strides in research have been made towards creating vaccinations and better understanding the way LASV infects cells fand the impact that the virus has on its host.
Introduction:
The Lassa Virus (commonly abbreviated LASV) is an old world zoonotic arenavirus. Its Baltimore Classification (a system that groups viruses based on their genome and replication method) is Class V, meaning that LASV has negative single stranded RNA that is translated directly into mRNA . Lassa virus is also a member of the arenaviridae family. According to Kehinde (2016), it infects between 100,000 and 500,000 people each year in West Africa. I researched scientific journal articles using the Web of Science database, and mainly examined the entry of the LASV virus into host cells and the possibility of a new method of vaccinating against the LASV.
Structurally, LASV’s nucleic acid genome is enveloped in a lipid bilayer. This genome consists of single-stranded negative RNA. The virus’s envelope is covered by a Class I trimeric glycoprotein spike complex. These spikes allow for function in unfavorable conditions, including more acidic pH’s that can be present in living biological cells. There are also two signal molecules, alpha and beta dystroglycan, present on the envelope’s surface. Genomically, the virus is structured in a bi-segmented fashion, one of which is called the Large (L) segment and the other the Small (S) segment. There are a total of four viral proteins, with each the L and S segments each coding for two proteins. The L segment codes for RNA polymerase and small zinc binding protein, while the S segment codes for nucleoprotein and glycoprotein precursors. These four proteins in combination work to regulate transcription and translation and assist in entry into the host cells.
Once within the body, the virus locates an antigen-producing host cell by binding to the α-dystroglycan primary cell receptor. The host cells are then triggered to ingest the virus through macropinocytosis. Once within the cell, the virus travels to an endosomal compartment. (Add about how fusion is triggered and release occurs?)
The virus is able to spread quickly in part due to its rapid replication process. The first step of LASV’s replication is the transcription of the mRNA of the negative genome. The positive genome then makes viral complementary RNA copies of itself. From these identical copies, mRNA is synthesized. The newly synthesized mRNA is then translated in order to synthesize Glycoprotein precursors as well as the zinc binding protein. The last proteins to be produced are the spike proteins that surround the enveloped genome. Yun et al (2012) said(REPLACE SAID) that this process utilizes temporal control, which benefits the virus by delaying the production of the identifying spike proteins until the very end. This allows the virus to complete its reproduction cycle within the cell before the spike proteins that signal that the virus is non-self are added, thereby delaying the immune response of the host cell.
After the spike proteins are produced and the virus is fully formed, the LASV seeks out an antigen- producing host cell, which it is able to enter through receptor mediated macropinocytosis (binding to the alpha-dystroglycan primary cell receptor). The host cell is then triggered to ingest the virus. Once within the cell, the virus travels to an endosomal compartment. This vesicle containing the virus then fuses with a late endosome. The low pH of the late endosome triggers the viral ribonucleoprotein to be released into the cytoplasm, where all the translation, transcription, and genomic replication occur. They then travel to the cell membrane, where the virions are assembled and the newly formed viruses bud off in a newly formed envelope. Lassa virus does little to no damage to the host cell as it reproduces, according to Peters et al (1989). (More recent source plz)
As soon as the virus is recognized in the body after infection, the immune system begins to produce Immunoglobulin G (IgG) and Immunoglobulin M (IgM) antibodies. These antibodies are often not a powerful enough of an attack on the virus, and the antibodies are unable to eradicate the virus from the host. The spike proteins trigger the immune response (HOW) , including the activation of T lymphocytes are thought to be instrumental in the survival of healthy host cells. This is because the T cells restrict the reproduction/replication of the virus and keep it from spreading throughout the host to as high a degree. In any subsequent infections by the same LASV virus, memory CD4 cells respond very strongly to the virus’s stimulation. This is an indication that the immune response is central to recovery from the virus. In order for the host to recover fully and in an efficient manner, an early diagnosis is essential (WHY). However, since the LASV’s symptoms present in a way similar to many other illnesses, its difficult to identify. Additionally, since the virus can only be identified through laboratory testing, areas with limited access to laboratories may have an increased amount of more serious cases.
The most common host for the Lassa virus is the African Soft-furred Rat (Mastomys natalensis). Kehinde (2016) states that in regions where the virus is endemic, up to thirty percent of the mastomys natalensis population is infected with LASV. Contact with the feces, urine, saliva, or other bodily fluids of an infected rodent by way of inhalation or ingestion can cause the transmission of the virus. This same direct contact with the bodily fluids of any infected person may also transmit it. After initial contact with the host, it may take up to three weeks for the virus to fully develop and for the symptoms to present themselves (CITE). The symptoms caused by viral infection by the LASV are common to the symptoms presented by many other acute viral hemorrhagic illness. These symptoms include abdominal and chest pain, cough, diarrhea, fever, headache, malaise, myalgia, nausea, sore throat, and vomiting. Many of the hemorrhagic illnesses that are present or endemic in many African countries share these same symptoms, making an initial diagnosis difficult. For the diagnosis to be confirmed, laboratory testing is required. This means that in areas where access to laboratories is limited, specific diagnoses may not always occur. According to Kehinde (2016), other symptoms may also arise as the disease progresses, including hemorrhaging, neurological problems, hearing loss, tremors, and encephalitis.
The Lassa Virus is not immediately detectable, nor are its symptoms immediately present. The virus can be incubated within the body for 7 to 21 days, after which the infection occurs. After this occurs, it generally takes 3-4 days before the virus is detectable using laboratory testing. However, since the LASV’s symptoms present in a way similar to many other illnesses, its difficult to identify. Additionally, since the virus can only be identified through laboratory testing, areas with limited access to laboratories may have an increased amount of more serious cases. In typical cases, patients begin to recover after 8 to 10 days. In the most severe of cases, deterioration of the patient’s condition occurs rapidly somewhere between the 6th and 10th days.
Research has shown that there are distinctions between the Lassa virus and other old world arenaviruses. In general, enveloped viruses are able to enter their host cells through interaction with cellular receptors (α-dystroglycan in the case of the Lassa virus) triggering macropinocytosis by the host cell, and membrane fusion with a late endosome whose low pH provokes the release of the viral ribonucleoprotein into the host cell’s cytoplasm. Here transcription, translation, and the replication of the viral ribonucleoprotein as a whole occurs. Researchers Israeli et al. (2017) discovered that in the case of the Lassa virus, the virus binds to the intracellular receptor LAMP1, as well, a different entry process than previously thought. Structurally, as mentioned previously, LASV has a glycoprotein spike complex, like many viruses, that is described as trimeric. This is made up of a structured signal protein, the GP1 receptor-binding molecule, and the GP2 trans-membrane module. Israeli H. et al (2017) demonstrated that three histidines on GP1 are essential when binding with LAMP1, indicating that LAMP1 and GP1’s interactions are an important part of the cell entry process of the Lassa virus, a concept that was not previously acknowledged. Based on these findings, the same researchers expected that other Old World arenaviruses would share this same process of entry. To prove or disprove this, they produced the GP1 receptor binding molecule, then introduced to a variety of isolated viruses in order to see whether binding and entry into the host cell occurred. The results of this are detailed in the Results section.
More recent research has also been done on a vaccination for the Lassa fever, as vaccinations are a powerful technique for preventing disease. In the past, there have been no effective LASV vaccines. Kainulainen et al. (2018) have recently conducted research in guinea pigs that introduces a potential vaccine for eventual human use. This vaccine introduces Lassa Virus Replicon Particles (VRPs) that lack the glycoprotein gene and a cell line with the glycoprotein products. Guinea pigs were injected subcutaneously with either 5×10^5 FFUs of wild-type VRPs (WT-WT VRPs), wild type VRPs with an inactivated S-segment encoded nucleoprotein exonuclease (WT-Exo(N) VRPs), or with Dulbecco’s modified Eagle’s medium (the control). No adverse clinical effects were found post vaccination. From this, Kainulainen et al. (2018) concludes that the VRPs did not cause Lassa fever in the guinea pig population. 4 weeks later, after blood testing, each of the guinea pigs was injected with 1×10^4 FFUs of LASV Josiah and then observed for 42 days. The results of this are also included below.
Results:
A pull down assay, where the presence of LAMP1 could be discerned using the anti-LAMP1 antibody, was used to test this. As shown in the table to the left, it was found that LAMP1 was present with LASV (Lassa Virus), but not MORV (Moriche Virus) or LCMV (Lymphocytic Choriomeningitis Virus).
Furthermore, for the vaccinated guinea pigs, the survival rates of each group is visible in the graph below (Kainulainen et al. (2018)). The guinea pigs injected with the control/Dulbecco’s modified Eagle’s medium began to experience elevated temperatures as well as weight loss after day 11. By day 23, all of these animals had died. All the animals vaccinated with either WT-WT VRPs or WT-Exo(N) VRPs not only survived the infection, but also did not experience any of the adverse symptoms that the control group did. After the 42 days, the tissue samples of the guinea pigs in the control group were found to contain LASV RNA in every case. However, the guinea pigs vaccinated with WT-WT VRPs or WT-Exo(N) VRPs had no tissue containing LASV RNA. ELISA was used to determine the presence of IgG immunoglobulin in each group before and after the experiment. Both the control group and the group of WT-WT VRPs lacked IgG ELISA titers either before or after the experiment took place. All of the guinea pigs in the WT-Exo(N) VRPs group developed IgG ELISA titers against the LASV by the 42nd day. This is visible in the graph to the left (Kainulainen et al. (2018)), which clearly shows that the subjects vaccinated with WT-Exo(N) VRPs were the only group, pre or post experiment, to develop IgG immunoglobulin of any kind. The significance of this is explained in the Conclusions section
Conclusions:
This indicates that LAMP1 is essential for Lassa virus, but not for either Moriche Virus of Lymphocytic Choriomeningitis Virus. The study by Israeli et al. (2017) indicates that MORV and LCMV are unable to bind with human LAMP1. The differentiation between LASV and other old world arenaviruses are also important in that their interactions are not as similar as we previously thought and that their differences in binding strategy may translate to other differences in the way they replicate or infect cells. (WHY? HOW?)
The results of the Kainulainen et al. (2018) study show that the immunoglobulin response caused by the vaccine could have long-lasting effects on the immunity of the vaccinated being to the LASV. If a vaccine is able to produce a long term effect/immune response without having any negative reactions in the vaccinated subject, as demonstrated in this study, then it is a viable option for a future vaccine. Though this experiment was conducted with guinea pigs, the results give hope that the use of the wild type VRPs with an inactivated S-segment encoded nucleoprotein exonuclease could, with more thorough research and testing, have the same effect in humans. There have been many past attempts to develop a vaccine to the Lassa virus, but none thus far have been successful for humans. Previous studies have successfully vaccinated subjects in animal models, including the use of live attenuated arenaviruses as a vaccine, the use of other viruses as vectors, and recombinant vaccinia viruses. However, these models have yet to be approved, and we cannot be sure of whether or not they would be effective in humans yet. The Lassa VRP approach described by Kainulainen et al. (2018), seems like it too would be a good option due to its scalable nature, as well as its stability, safety, and efficacy.
A better understanding of the Lassa Fever and a vaccine for the virus are both things we should be striving for in the future. Additionally , the discovery of a form of testing that doesn’t require a laboratory would be very beneficial in providing a prompt diagnosis and allowing for earlier intervention.