Uci’s ibc viral Vector Table




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UCI’s IBC Viral Vector Table
Recombination events or contamination from wildtype virus can result in the presence of replication competent virus (RCV) in a population of replication deficient viral vectors. (see vector table). Note that the IBC must review each individual project involving viral vectors to determine the appropriate biosafety level prior to initiating work with the vector. The biosafety levels listed below apply to replication incompetent vector systems only for in vitro (during production in tissue culture) and in vivo (rodents only, where all experiments would be terminal). In all cases, additional biosafety precautions may be recommended. The IBC always recommends testing for replication competent virus (RCV) in the vector stock. However, prior RCV testing of vector stock is required in order to house injected animals at Animal Biosafety Level-1 (ABSL-1) as indicated below.

For MSDS information click on highlighted vector


Viral Vector

Risk Group

Biosafety Level

Special Requirements for ABSL

Additional Precautions

Example Test Methods for Replication Competent Virus

in vitro

in vivo

(rodents)

Adenovirus 40 and 41



Adenovirus type 1, 2, 3, 4, 5 and 7


2
Hazards:

Droplet, aerosol, injection



BSL-2

ABSL-1 w/ special precautions1

Animals may be housed at ABSL-11 if vector stock is tested for RCV and found to be negative before injection
SOP’s and Animal Safety Form on door required.

All work done in a Biosafety Cabinet (BSC), Negative pressure lab preferred; SOP’s developed, Biological Hazard sign.
Transport specimens in a secondary, labeled, sealed container.
All waste picked up by EH&S

Vector stocks may be tested for RCV by PCR for E1a prior to use. The vector stock should be tested at a limit of sensitivity of 1 in 106 virus particles compared to a known positive control and the results of the test must be available upon request


For more vector information: http://medicine.ucsd.edu/gt/Adenovirus.html

ABSL-2


Without RCV testing of vector stock, or if stock tests positive, animals must be housed at ABSL-2
SOP’s and Animal Safety Form on door required.

Special Handling of bedding and cages for 48 hours post injection of animals. Incineration or autoclaving of bedding. Incineration of carcasses.



All work done in a Biosafety Cabinet, Negative pressure lab preferred; SOP’s developed, Biological Hazard sign.
Transport specimens in a secondary, labeled, sealed container
All waste picked up by EH&S

AAV (w/ adenovirus helper)

2

BSL-2

ABSL-1 w/ special precautions1



Animals may be housed at ABSL-11 if vector stock is tested for RCV (adenovirus) and found to be negative before injection. Special Handling of bedding and cages for 48 hours post injection of animals. Incineration or autoclaving of bedding. Incineration of carcasses.

All work done in a Biosafety Cabinet, Negative pressure lab preferred; SOP’s developed Biological Hazard sign.
Transport specimens in a secondary, labeled, sealed container
All waste picked up by EH&S

AAV vectors generated with adenovirus may be tested for the presence of replication competent adenovirus after heat inactivation at 56C for15 minutes.

<1 PFU of RCV/106 PFU recombinant virus

For more vector information http://medicine.ucsd.edu/gt/AAV.html

AAV (helper-free)

1

BSL-1

ABSL-1 w/ special precautions1



Animals may be housed at ABSL-1 with special precautions1 . Special Handling of bedding and cages for 48 hours post injection of animals. Incineration or autoclaving of bedding. Incineration of carcasses.




Not required

Baculovirus

1

BSL-1

ABSL-1 w/ special precautions1

Animals may be housed at ABSL-11 if vector stock is tested for RCV and found to be negative before injection


Use minimum requirement for Adenovirus.
All work done in a Biosafety Cabinet, Negative pressure lab preferred; SOP’s developed, Biological Hazard sign.
Transport specimens in a secondary, labeled, sealed container.
All waste picked up by EH&S




Herpesvirus type 1, 2; Herpes simplex virus (HSV)

2

BSL-2

ABSL-2

Animals should be housed at ABSL-2.
Filter top cages maybe required




Herpesvirus generated using amplicons must be tested for RCV by a plaque assay prior to approval for use at BSL-1 and ABSL-1. The vector stock should be tested at a limit of sensitivity of 1 infectious unit per milliliter and the test should include a known positive control. Herpesvirus vectors based on attenuated herpesvirus must always be handled at a BSL-2 level.






















Murine Retrovirus-

Ecotropic

1

BSL-1

ABSL-1 w/ special precautions1

Animals may be housed at ABSL-1 with special precautions1




Not required

Murine

Retrovirus-

Amphotropic (or VSV-G pseudotyped)

2

BSL-2

ABSL-1 w/ special precautions1

Animals may be housed at ABSL-11 if vector stock is tested for replication-competent virus (RCV) before injection




Retrovirus vector inoculums may be tested by amplifying any RCV present in permissive cell line (i.e. Mus dunni), and then screening by appropriate RCR detection assay (i.e. PG-4 S+L- assay, or the “marker rescue assay”*). The vector stock or producer line should be tested at a limit of sensitivity of 1 infectious unit per milliliter. 0 RCV in 106 infectious units

ABSL-2

If insert codes for a toxin or oncogene2, or if stock is not tested for RCV, animals must be housed at ABSL-2




Lentivirus
(3rd generation)


3

BSL-2

ABSL-1 w/ special precautions1



Animals may be housed at ABSL-1 if vector stock is tested for RCV and found to be negative before injection. Only one RCV test per construct is necessary. Subsequent preparations of virus from identical plasmids (identical insert, etc.) do not need to be tested.

-Needle protective devices should be used for injection procedures when possible

-Sealed rotor heads and/or canisters should be used for centrifugation and opened only in a biosafety cabinet

-Use only ultracentrifuges fitted with HEPA filters






Lentivirus vector stocks may be tested for RCV by serial transfer and ELISA assay for p24 antigen. The vector stock should be tested at a limit of sensitivity of 1 infectious unit per milliliter, and the test should include a known positive control.

Undetectable

p24 ELISA assay is preferred

(sensitivity depends on assay)



ABSL-2 w/ special precautions1

Without RCV testing of vector stock, or if stock tests positive, animals must be housed at ABSL-2. Additionally, if non-rodent cells are injected into animals, they must be kept at ABSL-2.

-Needle protective devices should be used for injection procedures when possible

-Sealed rotor heads and/or canisters should be used for centrifugation and opened only in a biosafety cabinet

-Use only ultracentrifuges fitted with HEPA filters






Retrovirus

2

BSL

1-2



ABSL-2

Work with amphotrophic viruses must be screened for replication competence before injection of animals (see Example methods);

All work done in a Biosafety Cabinet, full face protection if working outside a BSC.

Used engineered sharps.
Transport specimens in a secondary, labeled, sealed container.
All waste picked up by EH&S
Pseudotyping vectors often results in a higher biosafety level.

Retrovirus vector inoculums may be tested by amplifying any RCV present in permissive cell line (i.e. Mus dunni), and then screening by appropriate RCR detection assay (i.e. PG-4 S+L- assay, or the “marker rescue assay”*). The vector stock or producer line should be tested at a limit of sensitivity of 1 infectious unit per milliliter. 0 RCV in 106 infectious units

Sindbis virus

2

BSL-2

ABSl-2

Under review







Vaccinia

2

BSL-2

ABSL-2+

At UCI


-Personnel should be offered vaccination

Note: vaccination is not necessary for certain “highly attenuated” poxvirus strains (e.g. MVA, NYVAC, ALVAC, OR TROVAC which replicate poorly or not at all in human cells.

-Animals must be housed at ABSL-2+

-Needle protective devices should be used for injection procedures when possible

-Sealed rotor heads and/or canisters should be used for centrifugation and opened only in a biosafety cabinet

-Use only ultracentrifuges fitted with HEPA filters







Not applicable since used as replicating vector

Footnotes:

1ABSL-1 special precautions must be followed including: handling infected animals last, changing gloves after handling, clearly labeling cages and notifying ULAR prior to moving animals into room

2If not known, but can demonstrate that no oncogenic properties exist in cell culture, okay for ABSL-1 housing with testing

References:

Retrovirus



Wilson, C.A., Ng, T. H., and Miller, A. E., 1997. Evaluation of recommendations for replication-competent retrovirus testing associated with use of retroviral vectors. Human Gene Therapy, 8(7): 869-874.
Forestell, S.P., Nando, J. S., Bohnlein, E., and Rigg, R. J. 1996. Improved detection of replication-competent retrovirus. J Virol Methods 60: 171-178

Lentivirus


Reference for serial transfer and p24 ELISA assay: Dull T, Zufferey R, Kelly M, Mandel RJ, Nguyen M, Trono D, Naldini L. 1998. A third-generation lentivirus vector with a conditional packaging system. J Virol 72: 8463-8471.

Adeno Associated Virus

Reference for a RCV assay: Hehir KM, Armentano D, Cardoza LM, et a1. 1996. Molecular characterization of replication-competent variants of adenovirus vectors and genome modifications to prevent their occurrence. J Virol 70:8459-8467.


Adenovirus

Reference for E1a PCR assay: Zhang WW, Kock PE, Roth JA. 1995. Detection of wild-type contamination in a recombinant adenoviral preparation by PCR. Biotechniques 18: 444-447.



Herpesvirus
Reference for a plaque assay: Strathdee CA, McLeod MR. 2000. A modular set of helper-dependent herpes simplex virus expression vectors. Mol Ther 5:479-485.

Document from Stanford University:

http://www.stanford.edu/dept/EHS/prod/researchlab/bio/docs/Working_with_Viral_Vectors.pdf

Viral Vector Overview


Introduction: viruses and viral vectors have become a staple of the molecular biology community. As such, it is important for users to understand the origins of these tools and potential implications of their use. Sections for each virus contain information on virology, clinical features, epidemiology, treatment, laboratory hazards, Personnel Protective Equipment (PPE), disinfection, and use with animals.

General comment on containment: Suggested biosafety containment levels are provided for each vector system. Use of a higher-level containment facility may be required in some cases, depending on the specific properties of the vector and/or insert. Special care should be given to the design and handling of virus vectors containing genes that make growth-regulating products, products released into the circulation, products that may have a general effect on the host-immune system (see Viral Vector chart for more information). (1) Work with viral vectors that are classified as BSL-1 does not require Biosafety – common examples are Baculovirus and AAV (when oncogenes or toxins are not cloned into the vectors) and ecotropic MMLV. Work with BSL-2 or 3 agents require Biosafety approval prior to start of work. Additional approval from A-PLAC is required for research involving BSL-2/3 viral vectors and animals.

Click on link at end of each section for additional virus specific information,




    1. Adenovirus: Adenoviruses are infectious human viruses, which often cause mild respiratory illness, pink eye or gastroenteritis. Rare cases of severe disease can occur, and its use as a genetic vector therefore requires the use of adequate containment equipment and practices. Biosafety Level 2 (BL2) is appropriate for many constructs. Particular care should be given to vectors containing genes that make products that may be similar to products made by the deleted adenovirus genes. Additional Adenovirus information




    1. Adeno-associated virus (AAV): These are infectious human viruses with no known disease association. Some AAV types are common in the general population, and these viruses have the ability to integrate into the host chromosome. The NIH Guidelines (Appendix B) state that "adeno-associated virus (AAV) types 1 through 4, and recombinant AAV constructs, in which the transgene does not encode either a potentially tumorigenic gene product or a toxin molecule and are produced in the absence of a helper virus" can in most cases be handled at biosafety level 1 (BL1). This level of containment made is modified by other considerations (see above). Additional AAV information




    1. Epstein-Barr Virus (EBV): Epstein-Barr virus, frequently referred to as EBV, is a member of the herpesvirus family and one of the most common human viruses. The virus occurs worldwide, and most people become infected with EBV sometime during their lives. In the United States, as many as 95% of adults between 35 and 40 years of age have been infected. Infants become susceptible to EBV as soon as maternal antibody protection (present at birth) disappears. Many children become infected with EBV, and these infections usually cause no symptoms or are indistinguishable from the other mild, brief illnesses of childhood. In the United States and in other developed countries, many persons are not infected with EBV in their childhood years. When infection with EBV occurs during adolescence or young adulthood, it causes infectious mononucleosis 35% to 50% of the time. EBV also establishes a lifelong dormant infection in some cells of the body's immune system. A late event in a very few carriers of this virus is the emergence of Burkitt's lymphoma and nasopharyngeal carcinoma. EBV is a transforming virus and is often used to produce immortalized cell lines. BSL-2 is appropriate for most experiments. Additional EBV information




    1. Herpesvirus: Herpesviruses include infectious human viruses such as herpes simplex virus type-1 (HSV-1), which is the most commonly used vector system. HSV-1 is common in the general population, but can cause encephalitis in rare cases; its utility as a vector system stems from its broad host cell range, ability to transduce neurons, and its large insert capacity. Biosafety Level 2 (BL2) is appropriate for many constructs. Additional Herpesvirus information



    1. Retrovirus: These are infectious viruses which can integrate into transduced cells with high frequency, and which may have oncogenic potential in their natural hosts. Retrovirus vector systems are typically based on murine viruses - most commonly, these systems include ecotropic viruses (which can infect only murine cells), amphotropic viruses (which can infect human cells) or pseudotyped viruses, when vector particles express glycoproteins (GPs) derived from other enveloped viruses (which can also infect human cells). The most common GP currently used is VSV-g, however there are newer pseudotypes being derived from viruses such as measles (Rubeola), Ebola and Marburg. Pseudotyping vectors often results in a higher Biosafety level. Containment for vectors with the ability to infect human cells (amphotropic) will usually be recommended at BL2, whereas for ecotropic vectors with no ability to infect human cells, BL1 containment may be appropriate.


A. MMLV: The host range of recombinant MoMuLV vectors is dependent on the specificity of the viral envelope. The ecotropic env gene produces particles that infect only rodent cells. Amphotropic env gene allows infection of murine and nonmurine cells, including human cells. VSV-G envelope allows infection in a wide range of mammalian and non-mammalian cells. Biosafety Level 2 (BL2) is appropriate for many constructs, while higher levels may be required depending upon the construct. Additional MMVL information
B. Lentivirus: Lentiviruses are a subset of retroviruses, with the ability to integrate into host chromosomes, and to infect non-dividing cells. These viruses can cause severe immunologic and neurologic disease in their natural hosts. Lentivirus vector systems can include viruses of non-human origin (feline immunodeficiency virus, equine infectious anemia virus) as well as simian viruses (simian immunodeficiency virus) and human viruses (HIV). The more recent generation vectors have been designed to sufficiently diminish the possibility for recombination to occur resulting in a wildtype- potentially infectious virus. Typical lentivirus vectors are packaged using pseudotyped enveloped proteins. The most common envelope protein used for this purpose is from vesicular stomatitis virus (VSV). It is usually recommended that work with non-human lentiviruses that are incapable of establishing productive infections in humans be conducted at BL-2. Work with simian or human lentiviruses (SIV, HIV) is typically conducted at a higher containment level. Additional Lentivirus information

6. Poxvirus: Poxvirus vectors include avian viruses (avipox vectors) such as NYVAC and ALVAC, which cannot establish productive infections in humans, as well as mammalian poxviruses, which can productively infect humans -such as vaccinia virus and modified vaccinia viruses (MVA). Poxviruses are highly stable, and vaccinia virus can cause severe infections in immunocompromised persons, persons with certain underlying skin conditions, or pregnant women. Such individuals should not work with vaccinia virus. The use of BL2 is appropriate for many poxvirus and constructs. Additional Pox virus information
7.Baculovirus: Non-mammalian virus vectors that infect insects, they are very stable and may remain dormant in the environment for years before infecting insects. Work is mostly done at the BSL-1 level.


Adenovirus (2)




Virology: Adenoviruses are medium-sized (90-100 nm), nonenveloped icosohedral viruses containing double-stranded DNA. There are more than 49 immunologically distinct types (6 subgenera: A through F) that can cause human infections. Adenoviruses are unusually stable to chemical or physical agents and adverse pH conditions, allowing for prolonged survival outside of the body.
The adenovirus infection cycle can be clearly divided into two phases, which are separated by viral DNA replication. The first or "early" phase covers the entry of the virus into the host cell and the entry of the virus genome to the nucleus. The late genes are transcribed from the major late promoter. The “late” phase is involved in making gene products that are related to production and assembly of capsid proteins.

Virus packaged by transfecting HEK 293 cells with adenoviral-based vectors are capable of infecting human cells. These viral supernatants could, depending on the gene insert, contain potentially hazardous recombinant virus. Similar vectors have been approved for human gene therapy trials, attesting to their potential ability to express genes in vivo. For these reasons, due caution must be exercised in the production and handling of any recombinant adenovirus.


The probability of producing replication competent adenovirus (RCA), although low, increases with each successive amplification. RCA is produced when adenoviral DNA recombines with E1-containing genomic DNA in HEK 293 cells. It is suggested to use early amplification stocks when needed to produce additional quantities of adenovirus.
Clinical features: Adenoviruses most commonly cause respiratory illness; however, depending on the infecting serotype, they may also cause various other illnesses, such as gastroenteritis, conjunctivitis, cystitis, and rash-associated illnesses. Symptoms of respiratory illness caused by adenovirus infection range from the common cold syndrome to pneumonia, croup, and bronchitis. Patients with compromised immune systems are especially susceptible to severe complications of adenovirus infection that can cause more systemic diseases (e.g. hepatitis).
Epidemiology: Although epidemiologic characteristics of the adenoviruses vary by type, all are transmitted by direct contact, fecal-oral transmission, and occasionally waterborne transmission. Some types are capable of establishing persistent asymptomatic infections in tonsils, adenoids, and intestines of infected hosts, and shedding can occur for months or years. Some adenoviruses (e.g., serotypes 1, 2, 5, and 6) have been shown to be endemic in parts of the world where they have been studied, and infection is usually acquired during childhood. Other types cause sporadic infection and occasional outbreaks; for example, epidemic keratoconjunctivitis is associated with adenovirus serotypes 8, 19, and 37. Epidemics of febrile disease with conjunctivitis are associated with waterborne transmission of some adenovirus types. ARD is most often associated with adenovirus types 4 and 7 in the United States. Enteric adenoviruses 40 and 41 cause gastroenteritis, usually in children. For some adenovirus serotypes, the clinical spectrum of disease associated with infection varies depending on the site of infection; for example, infection with adenovirus 7 acquired by inhalation is associated with severe lower respiratory tract disease, whereas oral transmission of the virus typically causes no or mild disease.
Treatment: Most infections are mild and require no therapy or only symptomatic treatment. Because there is no virus-specific therapy, serious adenovirus illness can be managed only by treating symptoms and complications of the infection.
Laboratory hazards: Ingestion; droplet exposure of the mucous membrane

Laboratory Hazards PPE

Exposure of mucus membrane

(eyes, nose, mouth)



Use of safety goggles or full face shields.

Use of appropriate face mask



Injection

Use of safety needles; NEVER re-cap needle or remove needle from syringe

Aerosol inhalation

Use of appropriate respiratory protection

Direct contact with skin

Gloves, lab coat, closed shoes

The above PPE are often required IN ADDITION to working in a certified Biosafety Cabinet.


Susceptibility to disinfectants: Susceptible to 1% sodium hypochlorite, 2% glutaraldehyde, 0.25% sodium dodecyl sulfate
Use with Animals: BSL-2 housing for 48 hours post injection/exposure of animals.
Adenovirus MSDS

Adeno-associated virus(3)

Virology: Adeno-associated virus gets its name because it is often found in cells that are simultaneously infected with adenovirus. AAV are Parvoviridae; icosahedral, 20-25 nm in diameter; single stranded DNA genome with a protein capsid. AAV is dependent for replication on presence of wild type adenovirus or herpesvirus; in the absence of these helper viruses, AAV will stably integrate into the host cell genome. Co-infection with helper virus triggers a lytic cycle as do some agents which appropriately perturb host cells. Wild type AAV integrates preferentially into human chromosome 19q13.3-qter; recombinant vectors lose this specificity and appear to integrate randomly, thereby posing a theoretical risk of insertional mutagenesis
Clinical features: No known pathology for wild type AAV serotype 2.
Epidemiology: Not documented definitively. Infection apparently via mouth, esophageal, or intestinal mucosa.
Treatment: no specific treatment.
Laboratory hazards: Ingestion, droplet exposure of the mucous membrane, direct injection.
Laboratory Hazards PPE

Exposure of mucus membrane

(eyes, nose, mouth)



Use of safety goggles or full face shields.

Use of appropriate face mask



Injection

Use of safety needles; NEVER re-cap needle or remove needle from syringe

Aerosol inhalation

Use of appropriate respiratory protection

Direct contact with skin

Gloves, lab coat, closed shoes

The above PPE are often required IN ADDITION to working in a certified Biosafety Cabinet.


Susceptibility to disinfectants: Susceptible to 1% sodium hypochlorite, 2% glutaraldehyde, 0.25% sodium dodecyl sulfate

Use with Animals: BL1 housing.

Epstein-Barr Virus(4)

Virology: Double-stranded linear DNA, 120-150 nm diameter, enveloped, icosahedral; types A and B; Herpesviridae (Gammaherpesvirinae). Epstein-Barr virus (EBV), a ubiquitous B-lymphotrophic herpesvirus, has been found in the tumor cells of a heterogeneous group of malignancies (Burkitt's lymphoma, lymphomas associated with immunosuppression, other non-Hodgkin's lymphomas, Hodgkin's disease, nasopharyngeal carcinoma (NPC), gastric adenocarcinoma, lymphoepithelioma-like carcinomas, and immunodeficiency-related leiomyosarcoma). EBV is a transforming virus and can immortalize B-cells and cause lymphoma in various animal models.
Clinical Features: Infectious mononucleosis - acute viral syndrome with fever, sore throat, splenomegaly and lymphadenopathy; one to several weeks, rarely fatal/ Burkitt's lymphoma - monoclonal tumor of B cells, usually involving children, jaw involvement is common; AIDS patients( 25% -30% are EBV related) / Nasopharyngeal carcinoma - malignant tumor of epithelial cells of the nasopharynx involving adults between 20 and 40 years
Epidemiology: EBV infects 80 - 90% of all adults world wide; mononucleosis is common in early childhood worldwide, typical disease occurs in developed countries mainly in young adults; Burkitt's tumor is found worldwide but hyperendemic in highly malarial areas such as tropical Africa; carcinoma is worldwide but highest in Southeast Asia and China.

Transmission: Mononucleosis - person-to-person by oropharyngeal route via saliva, possible spread via blood transfusion (not important route); Burkitt's lymphoma - primary infection occurs early in life or involves immunosuppression and reactivation of EBV later, malaria an important co-factor; NPC is associated with EBV infection in early life and reactivation later with epithelial invasion.
Treatment: No specific treatment

Laboratory hazards: Ingestion, accidental parenteral injection, droplet exposure of the mucous membranes, inhalation of concentrated aerosolized materials. Note that cell lines are often immortalized by transformation with EBV.
Laboratory Hazards PPE

Exposure of mucus membrane

(eyes, nose, mouth)



Use of safety goggles or full face shields.

Use of appropriate face mask



Injection

Use of safety needles; NEVER re-cap needle or remove needle from syringe

Aerosol inhalation

Use of appropriate respiratory protection

Direct contact with skin

Gloves, lab coat, closed shoes

The above PPE are often required IN ADDITION to working in a certified Biosafety Cabinet.


Susceptibility to disinfectants: Susceptible to disinfectants - 1% sodium hypochlorite, 70% ethanol, glutaraldehyde, formaldehyde
Use with Animals: BL2 housing.
Epstein-Barr virus MSDS

Herpesvirus
Virology: Herpesviridae, Alphavirinae, genus Simplexvirus; double-stranded linear DNA virus, icosahedral, lipid envelope, 110 - 200 nm diameter, HSV types 1 and 2 can be differentiated immunologically. Vectors derived from Herpes simplex virus (HSV) have some unique features. The vectors have a wide host range and cell tropism, infecting almost every cell type in most vertebrates that have been examined. In addition, the natural property of the virus to infect and establish latent infection indefinitely in post-mitotic neurons has generated substantial interest in using it to deliver therapeutic genes to the nervous system.
Clinical Features: Classic presentation of primary HSV-1 is herpes gingivostomatitis - oral mucosa, HSV 1 - primary infection is usually mild (10% of cases can be severe) and in early childhood; reactivation of latent infection results in fever blisters or cold sores, usually on the face and lips which crust and heal within a few days, may be CNS involvement (meningoencephalitis), 70% mortality rate if left untreated; causes about 2% of acute pharyngotonsillitis; Classic presentation of a primary HSV-2 infection is herpes genitalis, HSV 2 - genital herpes, sexually transmitted, associated with aseptic meningitis, vaginal delivery can cause risk to newborn, encephalitis and death; either HSV-1 and HSV-2 may infect the genital tract or oral mucosa.
Epidemiology: Type 1 - contact with saliva of carriers, infection of hands of health care personnel; Type 2 - usually by sexual contact; infected secretions from symptomatic or asymptomatic individuals. Virus may be secreted in saliva for up to 7 weeks after recovery and from genital lesions for 7-12 days: asymptomatic oral and genital infections, with transient viral shedding, are common; reactivation can be precipitated by over-exposure to sunlight, febrile, physical or emotional stress or foods and drugs, especially chemotherapy; HSV may be shed intermittently from mucosal sites for years, possibly life long.
HSV is spread by direct contact with epithelial or mucosal surfaces. Additionally, approximately 50% - 90% of adults possess antibodies to HSV type 1; 20% - 30% of adults possess antibodies to HSV type 2.This is a concern as reactivation from latency is not well understood. Infection by HSV vectors into latently infected cells could potentially reactivate the wild-type virus, or spontaneous reactivation of a latent infection could produce an environment where replication defective vectors could replicate.
Treatment: anti-viral drug therapy for symptoms.
Laboratory Hazards: Ingestion; accidental parenteral injection; droplet exposure of the mucous membranes of the eyes, nose, or mouth; inhalation of concentrated aerosolized materials
Laboratory Hazards PPE

Exposure of mucus membrane

(eyes, nose, mouth)



Use of safety goggles or full face shields.

Use of appropriate face mask



Injection

Use of safety needles; NEVER re-cap needle or remove needle from syringe

Aerosol inhalation

Use of appropriate respiratory protection

Direct contact with skin

Gloves, lab coat, closed shoes

The above PPE are often required IN ADDITION to working in a certified Biosafety Cabinet.


Susceptibility to disinfectants: Susceptible to common disinfectants - 1% sodium hypochlorite, iodine solutions containing ethanol, 70% ethanol, glutaraldehyde, formaldehyde
Use with Animals: BL2 housing.
Herpes simplex virus MSDS

Lentivirus(5)
Virology: The genus of the family Retroviridae consists of non-oncogenic retroviruses that produce multi-organ diseases characterized by long incubation periods and persistent infection. Five serogroups are recognized, reflecting the mammalian hosts with which they are associated. HIV-1 is the type species.

Bovine lentiviruses (e.g. Bovine immunodeficiency virus, Jembrana disease virus)

Equine lentiviruses (e.g. Equine infectious anemia virus)

Feline lentiviruses (e.g. Feline immunodeficiency virus)

Ovine/caprine lentivirus (e.g. Caprine arthritis-encephalitis virus, Ovine lentivirus, Visna virus)

Primate lentivirus group

Human immunodeficiency virus (HIV) types 1 - 3

Simian AIDS retrovirus SRV-1

Human T-cell lymphotropic virus type I and II

Simian immunodeficiency virus 
Most of the lentiviral vectors presently in use are HIV-derived vectors. The cis- and trans-acting factors of lentiviruses are often on separate plasmid vectors, with packaging being provided in trans. The vector constructs contain the viral cis elements, packaging sequences, the Rev response element (RRE), and a transgene (6).
Lentiviral Pseudotyping

Replacement of the HIV envelope glycoprotein with VSV-G provides a broad host-range for the vector and allows the viral particles to be concentrated by centrifugation.


Clinical Features(7) : In terms of the pathogenesis of lentivirus, some key properties are:

  1. Lentiviruses persist lifelong. This is a function both of their ability to integrate into the host chromosome and of their ability to evade host immunity. This ability to evade host immunity may be related both to the high mutation rates of these viruses, and to their ability to infect immune cells (macrophages, and in the case of HIV, T-cells).

  2. Lentiviruses have high mutation rates. Lentiviruses replicate, mutate and undergo selection by host immune responses.

  3. Infection proceeds through at least three stages.

(A) Initial (acute) lentivirus infection is associated with rapid viral replication and dissemination, which is often accompanied by a transient period of disease.

(B) This is followed by a latent period, during which the virus is brought under immune control and no disease occurs.

(C) High levels of viral replication then resume at some later time, resulting in disease.
Epidemiology: Transmitted from person to person through direct exposure to infected body fluids (blood, semen) sexual contact, sharing unclean needles etc.; transplacental transfer can occur
Treatment: Specific measures for the opportunistic diseases that result from AIDS; multidrug treatment for HIV
Laboratory Hazards: Direct contact with skin and mucous membranes of the eye, nose and mouth; accidental parenteral injection; ingestion; hazard of aerosols exposure unknown
Laboratory Hazards PPE

Exposure of mucus membrane

(eyes, nose, mouth)



Use of safety goggles or full face shields.

Use of appropriate face mask



Injection

Use of safety needles; NEVER re-cap needle or remove needle from syringe

Aerosol inhalation

Use of appropriate respiratory protection

Direct contact with skin

Gloves, lab coat, closed shoes

The above PPE are often required IN ADDITION to working in a certified Biosafety Cabinet.


Susceptibility to disinfectants: Susceptible to many disinfectants - 1% sodium hypochlorite, 2% glutaraldehyde, formaldehyde, ethanol

Use with Animals: BSL-2 housing for 48 hours post injection of animals.
Lenti virus/Human Immunodeficiency Virus MSDS
Moloney Murine Leukemia Virus (MoMuLV) (8)
Virology: Retroviridae; subfamily oncovirinae type C, enveloped, icosahedral core, virions 100 nm in diameter, diploid, single stranded, linear RNA genome. MoMuLV integrates into the host genome and is present in infected cells as a DNA provirus. Cell division is required for infection. Virus is not lytic.
Data suggests a pathogenic mechanism in which chronic productive retroviral infection allowed insertional mutagenesis leading to cell transformation and tumor formation. The nature of a transgene or other introduced genetic element may pose additional risk.
The host range of recombinant MoMuLV vectors is dependent on the specificity of the viral envelope. The ecotropic env gene produces particles which infect only rodent cells. The amphotropic env gene allows infection of rodent and non-rodent cells, including human cells. VSV-G envelope allows infection in a wide range of mammalian and non-mammalian cells.
Clinical features: None to date.
Epidemiology: MoMuLV infects only actively dividing cells. In mice, the virus is transmitted in the blood from infected mother to offspring. Transmission may also occur via germline infection. In vivo transduction in humans appears to require direct injection with amphotropic or pseudotyped virus.
Treatment: No recommended treatment.
Laboratory Hazards: Contact with feces or urine from infected animals for 72 hours post infection. Contact with tissues and body fluids of infected animals. Direct injection.
Laboratory Hazards PPE

Exposure of mucus membrane

(eyes, nose, mouth)



Use of safety goggles or full face shields.

Use of appropriate face mask



Injection

Use of safety needles; NEVER re-cap needle or remove needle from syringe

Aerosol inhalation

Use of appropriate respiratory protection

Direct contact with skin

Gloves, lab coat, closed shoes

The above PPE are often required IN ADDITION to working in a certified Biosafety Cabinet.


Susceptibility to disinfectants: Susceptible to many disinfectants - 1% sodium hypochlorite, 2% glutaraldehyde, formaldehyde, ethanol

Use with Animals: BL1 housing for ecotropic, BSL-2 for amphotropic or pseudotyped vector
Pox viruses/Vaccinia (9)

Virology: The poxviruses are the largest known DNA viruses and are distinguished from other viruses by their ability to replicate entirely in the cytoplasm of infected cells. Poxviruses do not require nuclear factors for replication and, thus, can replicate with little hindrance in enucleated cells. The core contains a 200-kilobase (kb), double-stranded DNA genome and is surrounded by a lipoprotein core membrane.
Recombinant Vaccinia vectors

Vaccinia virus can accept as much as 25 kb of foreign DNA, making it useful for expressing large eukaryotic and prokaryotic genes. Foreign genes are integrated stably into the viral genome, resulting in efficient replication and expression of biologically active molecules. Furthermore, posttranslational modifications (e.g., methylation, glycosylation) occur normally in the infected cells.


Vaccinia is used to generate live recombinant vaccines for the treatment of other illnesses. Modified versions of vaccinia virus have been developed for use as recombinant vaccines. The modified Ankara strain (MVA) of vaccinia virus was developed by repeated passage in a line of chick embryo fibroblasts. NYVAC is another attenuated form of the vaccinia virus that has been used in the construction of live vaccines. NYVAC has a deletion of 18 vaccinia virus genes that render it less pathogenic.
Clinical Features: Virus disease of skin induced by inoculation for the prevention of smallpox - vesicular or pustular lesion, area of induration or eythema surrounding a scab or ulcer at inoculation site; major complications encephalitis, progressive vaccinia (immunocompromised susceptible), eczema vaccinatum, fetal vaccinia; minor complications - generalized vaccinia with multiple lesions; auto-inoculation of mucous membranes or abraded skin, benign rash, secondary infections; complications are serious for those with eczema or who are immunocompromised
Epidemiology: Communicable to unvaccinated contacts via contact with mucosal membranes or cuts in skin.
Treatment: Vaccinia immune globulin and an antiviral medication may be of value in treating complications
Vaccination: Consultation is available to determine if vaccination with the Smallpox vaccine is appropriate for personnel using vaccinia.
Laboratory Hazards: Ingestion, parenteral injection, droplet or aerosol exposure of mucous membranes or broken skin with infectious fluids or tissues.
Laboratory Hazards PPE

Exposure of mucus membrane

(eyes, nose, mouth)



Use of safety goggles or full face shields.

Use of appropriate face mask



Injection

Use of safety needles; NEVER re-cap needle or remove needle from syringe

Aerosol inhalation

Use of appropriate respiratory protection

Direct contact with skin

Gloves, lab coat, closed shoes

The above PPE are often required IN ADDITION to working in a certified Biosafety Cabinet.


Susceptibility to disinfectants: Susceptible to 1% sodium hypochlorite, 2% glutaraldehyde, formaldehyde
Use with Animals: BL2+ housing.
Pox/Vaccinia virus MSDS

References




  1. http://www.hawaii.edu/ehso/bio/BSM_part21.htm




  1. http://www.cdc.gov/ncidod/dvrd/revb/respiratory/eadfeat.htm




  1. http://medicine.ucsd.edu/gt/AAV.html




  1. http://www.phac-aspc.gc.ca/msds-ftss/msds62e.html




  1. http://medical.webends.com/kw/Lentivirus




  1. http://www.unifr.ch/biochem/DREYER/LENTIVIRU1.htm




  1. http://biology.kenyon.edu/slonc/gene-web/Lentiviral/Lentivi2.html




  1. http://medicine.ucsd.edu/gt/MoMuLV.html




  1. http://www.emedicine.com/MED/topic2356.htm





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