Lassa Virus- An Overview

Lassa virus (LASV) is a single-stranded RNA virus that belongs to the family Arenaviridae, genus Mammarenavirus. It is one of the causative agents of viral hemorrhagic fever (VHF), a severe and often fatal disease that affects multiple organs and systems in humans. Lassa virus was first isolated and identified in 1969, after an outbreak of VHF among nurses in a hospital in Lassa, a town in northeastern Nigeria. Since then, Lassa virus has been recognized as an emerging and re-emerging pathogen that poses a significant public health threat in West Africa, where it is endemic in several countries, including Sierra Leone, Liberia, Guinea, and Nigeria. It is estimated that Lassa virus infects 100,000 to 300,000 people annually in this region, resulting in about 5,000 deaths. Lassa virus can also cause sporadic cases or outbreaks in travelers or health care workers who have been exposed to the virus in endemic areas or in laboratory settings.

Lassa virus is a zoonotic virus, meaning that it is transmitted to humans from animals. The natural reservoir of Lassa virus is a rodent of the genus Mastomys, commonly known as the multimammate rat. These rodents are widely distributed and abundant in West Africa, and can shed the virus in their urine and feces without showing any signs of illness. Humans can become infected with Lassa virus by direct or indirect contact with contaminated rodent excreta or materials, such as food or household items. Human-to-human transmission can also occur through contact with infected blood or body fluids of symptomatic patients or deceased persons. Nosocomial transmission, or transmission within health care facilities, is a major concern for Lassa virus infection, especially in resource-limited settings where infection prevention and control measures are inadequate.

The incubation period of Lassa virus infection ranges from 6 to 21 days. The clinical manifestations of Lassa virus infection are variable and nonspecific, making diagnosis challenging. About 80% of infected individuals have mild or no symptoms. However, about 20% of infected individuals develop severe disease that can affect multiple organs, such as the liver, spleen, kidneys, lungs, heart, and central nervous system. The common signs and symptoms of Lassa virus infection include fever, headache, malaise, sore throat, cough, chest pain, nausea, vomiting, diarrhea, abdominal pain, and facial swelling. In severe cases, patients may develop bleeding from various sites, such as the mouth, nose, gums, eyes, ears, vagina, or gastrointestinal tract. Other complications include shock, seizures, tremor, disorientation, coma, deafness (which may be permanent), hair loss (which may be temporary), and gait disturbance (which may be temporary). The case fatality rate of Lassa virus infection is about 1% overall but can reach up to 15% among hospitalized patients with severe disease.

The diagnosis of Lassa virus infection is based on laboratory testing of blood or tissue samples. Several methods are available for detecting the presence of the virus or its components (such as RNA or antigens) or the host immune response (such as antibodies) to the virus. These methods include reverse transcription polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), immunofluorescence assay (IFA), immunohistochemistry (IHC), and virus isolation by cell culture. However, these methods require specialized equipment and biosafety level 4 (BSL-4) laboratories that are not widely available in endemic areas. Therefore, rapid diagnostic tests (RDTs) that can be performed at the point of care with minimal resources are needed for timely diagnosis and management of Lassa virus infection.

The treatment of Lassa virus infection consists of supportive care and antiviral therapy. Supportive care includes hydration, electrolyte balance correction,

Lassa virus is a single-stranded RNA virus belonging to the family Arenaviridae. The virus has a spherical shape with an average diameter of 90–110 nm. The virus is enveloped in a lipid membrane with glycoprotein spikes protruding from the surface. The glycoproteins form T-shaped structures that extend 7–10 nm from the envelope and mediate the attachment and entry of the virus into host cells .

The structure of the Lassa virus spike complex was recently solved by Katz et al. (2022), who revealed that the virus binds to its cellular receptor, matriglycan, a linear carbohydrate that is present on alpha-dystroglycan. The authors also showed that the spike complex contains a stable signal peptide (SSP) that crosses the membrane once and helps to stabilize the spike in its native conformation.

The genome of Lassa virus consists of two segments of RNA, called the small (S) and large (L) segments. Each segment has two genes at opposite ends that do not overlap. The S segment encodes the nucleoprotein (NP) and the glycoprotein precursor (GPC), which is cleaved into GP1 and GP2 by a protease. The L segment encodes the zinc-binding protein (Z) and the RNA polymerase (L) .

The genome structure of Lassa virus is shown in the following figure:

 5` NP GPC 3`
S: UUUUUUUUUUUUUUUUUUUU
|||||||||||||||||||
AAAAAAAAAAAAAAAAAAA
L: 3` Z L 5`

Lassa virus has four lineages, with a strain variation of 27% in relation to their nucleotides and 15% in relation to their amino acids. Lineage I, II, and III are found in Nigeria, while lineage IV is found in Guinea, Liberia, and Sierra Leone .

Lassa virus has a single-stranded RNA genome that consists of two segments: the large (L) segment and the small (S) segment. Each segment has two genes that are encoded in opposite orientations and do not overlap. The L segment encodes the Z protein and the L protein, while the S segment encodes the glycoprotein precursor (GPC) and the nucleoprotein (NP). The Z protein is a small zinc-binding protein that regulates viral transcription and replication. The L protein is the RNA-dependent RNA polymerase that catalyzes viral RNA synthesis. The GPC is cleaved into two envelope glycoproteins, GP1 and GP2, that mediate viral attachment and entry into host cells. The NP is the major structural protein that forms the viral ribonucleoprotein complex with the genomic RNA.

The L segment is about 7.3 kilobases long and has a 5` untranslated region (UTR) of 55 nucleotides and a 3` UTR of 221 nucleotides. The Z gene is located at the 5` end of the L segment and has an open reading frame (ORF) of 276 nucleotides, encoding a protein of 92 amino acids. The L gene is located at the 3` end of the L segment and has an ORF of 6,633 nucleotides, encoding a protein of 2,211 amino acids.

The S segment is about 3.4 kilobases long and has a 5` UTR of 42 nucleotides and a 3` UTR of 63 nucleotides. The GPC gene is located at the 5` end of the S segment and has an ORF of 1,473 nucleotides, encoding a protein of 491 amino acids. The NP gene is located at the 3` end of the S segment and has an ORF of 1,710 nucleotides, encoding a protein of 570 amino acids.

Lassa virus belongs to the Old World arenaviruses and has four distinct lineages: lineage I, II, III, and IV. Lineage I is found in Nigeria, lineage II is found in Guinea, Liberia, and Sierra Leone, lineage III is found in Mali, and lineage IV is found in Ivory Coast. The four lineages differ by about 27% in their nucleotide sequences and by about 15% in their amino acid sequences.

The genome of Lassa virus has some unique features that distinguish it from other RNA viruses. For example, it has some double-stranded regions that form stem-loop structures in both segments. These structures are involved in viral transcription termination and initiation. Another feature is that Lassa virus uses a cap-snatching mechanism to acquire the cap structures from host mRNAs for its own mRNAs. This mechanism is mediated by the endonuclease activity of the L protein and the cap-binding activity of the NP protein.

The genome of Lassa virus is also prone to mutation and recombination, which contribute to its genetic diversity and evolution. Mutation rates vary among different genes and regions of the genome. Recombination events occur mainly between different strains or lineages of Lassa virus within the same host or during co-infection. Recombination can result in novel viral variants with altered antigenicity, virulence, or tropism.

The genome of Lassa virus is essential for its replication cycle, pathogenesis, and interaction with the host immune system. Understanding its structure, function, and diversity can provide insights into the molecular biology and epidemiology of this important human pathogen.

Lassa virus is endemic in parts of West Africa, including Sierra Leone, Liberia, Guinea and Nigeria. The virus was first isolated and identified as the cause of Lassa fever in 1969 in a small town called Lassa in northeastern Nigeria. The virus belongs to four lineages, three of which are found in Nigeria and one of which is distributed in Guinea, Liberia and Sierra Leone.

The natural reservoir of Lassa virus is the multimammate rat (Mastomys natalensis), which is widely distributed throughout West Africa. Humans can contract the virus through contact with the urine, feces or blood of infected rodents, or through ingestion of contaminated food or household items. The virus can also be transmitted from person to person through direct contact with the blood, urine, feces or other bodily secretions of an infected person. There is no evidence of airborne transmission between humans.

The annual number of Lassa fever cases in West Africa is estimated to range from 100,000 to 300,000, with about 5,000 deaths. However, surveillance for Lassa fever varies between locations and these estimates are likely to be underestimates. In some areas of Sierra Leone and Liberia, about 10-16% of people admitted to hospitals every year have Lassa fever, indicating the serious impact of the disease on the population of these regions.

Lassa fever is a seasonal disease that occurs more frequently during the dry season (November to April) than during the rainy season (May to October). This may be related to the breeding patterns and behavior of the multimammate rats. Lassa fever can affect people of all ages and both sexes, but pregnant women are particularly vulnerable to severe complications and death from the infection . The case fatality rate among pregnant women can reach 80% and the virus can cause spontaneous abortion, stillbirth or neonatal death .

Lassa fever is a public health threat that requires enhanced surveillance, prevention and control measures in West Africa. The World Health Organization (WHO) provides technical and operational support to countries affected by Lassa fever outbreaks and coordinates regional and international response efforts.

Lassa virus is a zoonotic disease, meaning that humans become infected from contact with infected animals. The animal reservoir, or host, of Lassa virus is a rodent of the genus Mastomys, commonly known as the “multimammate rat” . Mastomys rats infected with Lassa virus do not become ill, but they can shed the virus in their urine and faeces.

Transmission of Lassa virus to humans normally occurs through contamination of broken skin or mucous membranes via direct or indirect contact with infected rodent excreta on floors, home surfaces, in food or water. Transmission is also possible where rodents are caught and consumed as food. Contact with the virus may also occur when a person inhales tiny particles in the air contaminated with infected rodent excretions. This aerosol or airborne transmission may occur during cleaning activities, such as sweeping.

Direct contact with infected rodents is not the only way in which people are infected; person-to-person transmission may occur after exposure to virus in the blood, tissue, secretions, or excretions of a Lassa virus-infected individual. Casual contact (including skin-to-skin contact without exchange of body fluids) does not spread Lassa virus. Person-to-person transmission is common in health care settings (called nosocomial transmission) where proper personal protective equipment (PPE) is not available or not used . Lassa virus may be spread in contaminated medical equipment, such as reused needles.

The Lassa virus gains entry into the host cell by binding to the cell-surface receptor alpha-dystroglycan (alpha-DG), a protein that is involved in cell adhesion and signaling. The virus then enters the cell by endocytosis, a process that involves the formation of a vesicle around the virus particle and its transport into the cell. Inside the vesicle, the low pH triggers the fusion of the viral envelope with the vesicle membrane, releasing the viral ribonucleoprotein (RNP) complex into the cytoplasm. The RNP complex consists of the viral RNA genome and the viral proteins NP (nucleoprotein) and L (RNA polymerase).

The Lassa virus has a segmented, single-stranded RNA genome with negative polarity, meaning that it is complementary to the mRNA that is used for protein synthesis. The genome consists of two segments: the small (S) segment and the large (L) segment. The S segment encodes two proteins: NP and GP (glycoprotein precursor), which is cleaved into GP1 and GP2, the surface glycoproteins that form the spikes on the viral envelope. The L segment encodes two proteins: Z (zinc-binding protein), which regulates transcription and replication, and L, which is the RNA polymerase.

The first step of viral replication is transcription, which involves the synthesis of mRNA copies of the viral genome segments by the L polymerase. The L polymerase uses a cap-snatching mechanism to acquire a 5` cap structure from cellular mRNAs, which is essential for mRNA stability and translation. The cap-snatching mechanism involves the endonuclease activity of the L polymerase, which cleaves a short fragment from cellular mRNAs, and the cap-binding activity of NP, which binds to the cap structure. The L polymerase then uses this fragment as a primer to transcribe mRNA from the viral genome segments.

The second step of viral replication is translation, which involves the synthesis of viral proteins from the viral mRNAs by cellular ribosomes. The NP and L proteins are translated from the mRNAs transcribed from the genomic RNA segments (negative sense), while the GP and Z proteins are translated from the mRNAs transcribed from the antigenomic RNA segments (positive sense). The GP protein is post-translationally modified in the endoplasmic reticulum, where it is cleaved into GP1 and GP2 by cellular proteases. The GP1 and GP2 proteins form heterodimers that are transported to the Golgi apparatus and then to the plasma membrane, where they are incorporated into new virus particles.

The third step of viral replication is genome replication, which involves the synthesis of new viral genome segments by the L polymerase. The L polymerase uses both genomic and antigenomic RNA segments as templates to synthesize complementary RNA strands. The newly synthesized RNA strands then pair with their respective template strands to form double-stranded RNPs. The RNPs are then packaged into new virus particles along with NP, Z and L proteins.

The final step of viral replication is budding, which involves the release of new virus particles from the host cell by exocytosis. The budding process is mediated by Z protein, which interacts with both GP proteins on the plasma membrane and RNPs in the cytoplasm. The Z protein also recruits cellular factors that facilitate membrane curvature and scission. The new virus particles then exit the cell and infect new cells or hosts.

Lassa virus (LASV) is an Old World arenavirus that causes Lassa fever, a severe viral hemorrhagic fever in humans. The pathogenesis of LASV is not fully understood, but it involves multiple factors, such as viral replication, immune evasion, host response, and tissue damage.

Viral replication

LASV enters the host cell by binding to the cell-surface receptor alpha-dystroglycan (alpha-DG), which is widely expressed in various tissues . After endocytosis, the virus fuses with the endosomal membrane and releases its ribonucleoprotein (RNP) complex into the cytoplasm. The RNP consists of two segments of negative-sense RNA (S and L) and the viral proteins nucleoprotein (NP) and RNA polymerase (L). The viral RNA is transcribed and replicated by the L polymerase, using a cap-snatching mechanism to acquire the 5` cap from cellular mRNAs . The S segment encodes the glycoprotein precursor (GPC) and NP, while the L segment encodes the Z protein and L polymerase. The GPC is cleaved into two subunits, GP1 and GP2, which form the spikes on the viral envelope. The Z protein is a matrix protein that mediates virus assembly and budding .

LASV replicates mainly in dendritic cells (DCs) and macrophages, which are important for initiating innate and adaptive immune responses. However, LASV also infects other cell types, such as endothelial cells, hepatocytes, adrenal cells, and epithelial cells, leading to widespread tissue damage .

Immune evasion

LASV has evolved several strategies to evade the host immune system and establish persistent infection. One of these strategies is the expression of NP, which has anti-interferon (IFN) activity. NP inhibits the induction of type I IFN by blocking the signaling pathways of RIG-I-like receptors (RLRs) and Toll-like receptors (TLRs), which are sensors of viral RNA . NP also inhibits the activation of IRF-3 and IRF-7, transcription factors that induce IFN expression . Furthermore, NP antagonizes the antiviral effects of IFN by degrading STAT1, a key component of the IFN signaling pathway .

Another strategy of LASV to evade immune recognition is the downregulation of major histocompatibility complex (MHC) class I molecules on infected cells. MHC class I molecules present viral peptides to cytotoxic T lymphocytes (CTLs), which can kill infected cells. LASV reduces the expression of MHC class I molecules by interfering with their transport from the endoplasmic reticulum to the cell surface . This impairs the presentation of viral antigens and reduces the activation of CTLs.

Host response

Despite the immune evasion mechanisms of LASV, some infected individuals can mount an effective immune response and clear the virus. The host response to LASV involves both innate and adaptive immunity.

The innate immune response to LASV is mediated by natural killer (NK) cells, macrophages, neutrophils, and monocytes. These cells produce pro-inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), IL-8, IL-12, and IL-18, which activate other immune cells and induce inflammation . NK cells also lyse infected cells by releasing perforin and granzymes .

The adaptive immune response to LASV is mediated by B cells and T cells. B cells produce antibodies against LASV antigens, such as GPC, NP, Z protein, and L polymerase. Antibodies can neutralize the virus by blocking its attachment to alpha-DG or its fusion with endosomal membranes . Antibodies can also mediate antibody-dependent cellular cytotoxicity (ADCC) by recruiting NK cells or macrophages to kill infected cells . T cells can be divided into CD4+ T helper (Th) cells and CD8+ CTLs. Th cells secrete cytokines that help B cell differentiation and antibody production. CTLs recognize viral peptides presented by MHC class I molecules and kill infected cells by releasing perforin and granzymes .

Tissue damage

The tissue damage caused by LASV infection is mainly due to direct viral cytopathic effects and excessive inflammation. LASV infection induces apoptosis in various cell types, such as DCs, macrophages, endothelial cells, and hepatocytes . Apoptosis is triggered by the activation of caspases, which are proteases that execute the cell death program. LASV also induces necrosis in some cell types, such as adrenal cells and epithelial cells . Necrosis is a form of cell death that is characterized by membrane rupture and release of cellular contents, which can trigger inflammation and tissue injury.

The excessive inflammation induced by LASV infection can also contribute to tissue damage and organ failure. The pro-inflammatory cytokines produced by immune cells can cause vasodilation, increased vascular permeability, edema, hypotension, and shock . The cytokines can also activate the coagulation cascade, leading to disseminated intravascular coagulation (DIC), hemorrhage, and thrombosis . Moreover, the cytokines can induce tissue damage by activating cytotoxic cells, such as NK cells, neutrophils, and macrophages, which release reactive oxygen species (ROS) and proteases that degrade the extracellular matrix .

The clinical manifestations of Lassa virus infection vary widely, ranging from asymptomatic to severe and fatal. The incubation period of Lassa fever is typically 6 to 21 days, depending on the route of exposure and the viral load.

About 80% of Lassa fever cases are mild or subclinical, with symptoms such as slight fever, headache, general malaise, and weakness. These cases are often undiagnosed and may resolve spontaneously without specific treatment.

However, about 20% of Lassa fever cases progress to severe disease, characterized by hemorrhagic, respiratory, gastrointestinal, neurological, and renal manifestations. The onset of severe symptoms is usually gradual, but may be abrupt in some cases.

Some of the common signs and symptoms of severe Lassa fever include :

• Bleeding in the gums, nose, eyes, or elsewhere
• Difficulty breathing, cough, and chest pain
• Swollen airways and facial swelling
• Vomiting and diarrhea, both with blood
• Difficulty swallowing and sore throat
• Hepatitis and jaundice
• Abdominal pain and constipation
• Shock and low blood pressure
• Seizures, tremors, disorientation, and coma
• Hearing loss and deafness

The severity of Lassa fever may depend on several factors, such as the viral strain, the host immune response, the presence of underlying conditions, and the timeliness of treatment. The case-fatality rate of Lassa fever is estimated at 1% for all infections and 15% to 20% for hospitalized patients. The mortality rate is higher for pregnant women in their third trimester, with a 95% chance of fetal loss.

The diagnosis of Lassa fever is challenging due to the nonspecific and variable nature of the symptoms. Laboratory tests are required to confirm the infection and rule out other causes of febrile illness. The treatment of Lassa fever consists mainly of supportive care and antiviral therapy with ribavirin if initiated early in the course of the disease.

Lassa fever is a serious public health problem in West Africa, where it causes recurrent outbreaks and affects thousands of people every year. The prevention and control of Lassa fever depend on reducing the exposure to the rodent reservoirs and interrupting the human-to-human transmission through infection prevention and control measures.

Lassa virus infection can be difficult to diagnose clinically, as the signs and symptoms are varied and nonspecific. Therefore, laboratory tests are essential for confirming the diagnosis. The following tests can be used to detect Lassa virus or its components in blood or tissue samples :

• Reverse transcription-polymerase chain reaction (RT-PCR): This test can amplify and detect the viral RNA in the early stage of disease, usually within the first 10 days of symptom onset. RT-PCR is sensitive and specific, but it requires specialized equipment and trained personnel .
• Enzyme-linked immunosorbent serologic assays (ELISA): This test can measure the IgM and IgG antibodies against Lassa virus, as well as the viral antigen, in serum samples. ELISA is the most commonly used diagnostic test for Lassa fever, as it is relatively simple, inexpensive, and widely available. However, ELISA may not detect antibodies or antigen in the early phase of infection, and it may cross-react with other arenaviruses .
• Antigen detection tests: These tests can identify the viral antigen in urine or throat swabs using immunochromatographic strips or lateral flow devices. Antigen detection tests are rapid and easy to perform, but they have lower sensitivity and specificity than RT-PCR or ELISA.
• Virus isolation by cell culture: This test can grow and isolate the virus from blood or tissue samples using cell lines such as Vero E6 or BHK-21. Virus isolation by cell culture is the gold standard for diagnosing Lassa virus infection, as it provides direct evidence of viral presence and allows for further characterization and typing of the virus. However, this test is slow, labor-intensive, and requires a high-containment laboratory with biosafety level 4 (BSL-4) facilities .
• Immunohistochemistry: This test can detect the viral antigen in formalin-fixed tissue specimens using specific antibodies and colorimetric or fluorescent detection methods. Immunohistochemistry can be used to make a post-mortem diagnosis of Lassa fever in cases where blood or tissue samples are not available or suitable for other tests. However, this test also requires a high-containment laboratory with BSL-4 facilities.

The choice of diagnostic test depends on several factors, such as the availability of resources, the stage of disease, the type of sample, and the purpose of testing. Ideally, a combination of tests should be used to increase the accuracy and reliability of diagnosis. In addition, infection control measures should be followed to prevent exposure and transmission of Lassa virus during sample collection, handling, transport, and testing .

The treatment of Lassa fever is mainly based on the use of an antiviral drug called ribavirin, which has been shown to be effective if given early in the course of the illness . Ribavirin inhibits the replication of the Lassa virus by interfering with its RNA synthesis. The recommended dose of ribavirin for Lassa fever is 30 mg/kg intravenously (IV) as a loading dose, followed by 16 mg/kg IV every 6 hours for 4 days, and then 8 mg/kg IV every 8 hours for 6 days. Ribavirin can reduce the mortality rate of Lassa fever from about 50% to less than 10% if administered within the first 6 days of symptom onset .

However, ribavirin is not a specific treatment for Lassa fever and has some limitations and side effects. Ribavirin is not effective against all strains of Lassa virus and may not be available or affordable in some endemic areas. Ribavirin can also cause hemolytic anemia, teratogenicity, and mutagenicity, which may limit its use in pregnant women and children. Therefore, there is a need for more specific and safer treatments for Lassa fever.

In addition to ribavirin, patients with Lassa fever should also receive supportive care, which includes maintaining adequate fluid and electrolyte balance, oxygenation and blood pressure, as well as treating any secondary infections or complications . Supportive care can help prevent or reduce the severity of organ failure, shock, bleeding, and neurological manifestations that may occur in severe cases of Lassa fever .

Currently, there are no approved vaccines for Lassa fever, although several candidates are under development and testing . Vaccines could potentially prevent or reduce the transmission and morbidity of Lassa fever in endemic areas. However, the development of effective vaccines faces several challenges, such as the genetic diversity of Lassa virus strains, the lack of animal models that mimic human disease, and the ethical and logistical issues of conducting clinical trials in resource-limited settings .

Therefore, the treatment of Lassa fever remains a major public health challenge that requires more research and investment to develop novel therapeutics and vaccines. Until then, early diagnosis and prompt administration of ribavirin and supportive care are the mainstay of treatment for Lassa fever.

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