A review of post-exposure strategies for high-consequence viral pathogens in the laboratory

SUMMARY

Laboratory-acquired infections (LAIs), particularly those from high-risk viruses, pose significant threats to exposed individuals and to the general public. In this review, we evaluate the existing evidence for viral LAI prevention, including available vaccinations, post-exposure prophylaxis (PEP), and follow-up procedures following occupational exposure to Risk Group 3 and 4 viral infectious agents within clinical testing laboratories. This review provides guidance on the therapeutic options and follow-up, all essential for preparedness planning and timely management in the event of exposure.

INTRODUCTION

Laboratory-acquired infections (LAIs), particularly those from high-risk viruses, pose significant threats to exposed individuals and to the general public. Following the COVID-19 pandemic, the World Health Organization (WHO) adopted a list of priority pathogens, including many highly pathogenic viruses, with the intention to drive research and future pandemic preparedness. In addition, recent outbreaks of Sudan Ebola virus (SUDV), mpox, and avian influenza have resulted in increased diagnostic and research activities within higher biosafety level (BSL) facilities around the world (1, 2).

Despite significant efforts to improve and standardize laboratory biosafety procedures by the WHO Technical Advisory Group on Biosafety, significant challenges remain, particularly in under-resourced settings. The risk of LAI can vary significantly depending on the following: the techniques in handling viral preparations; individual safety preparation in the correct physical containment setting; and appropriate personal protective equipment (PPE). Handling of highly pathogenic virus in a concentrated form such as cell culture and performing aerosolizing or invasive procedures, such as centrifugation, vortexing, or venipuncture, are of particular concern and require preventative strategies. These include adequate staff training, PPE, and the appropriate facilities. Processing and handling of primary clinical samples can also pose a significant risk if these control measures are not in place to minimize the risk of exposure (3).

Given the complexity of the risk assessment process following accidental exposure, procedures for reporting and post-exposure management should be in place. These would assist both clinicians and laboratory staff in ensuring appropriate consideration is given to the prevention of LAIs. Despite this need, there is a paucity of formal consolidated guidelines considering LAIs with high-risk viral pathogens, largely due to lack of research in informing exposure risks and prophylaxis efficacy. Such challenges may be primarily attributed to the sporadic nature of human viral outbreaks (4), though difficulties in conducting animal challenge studies, especially non-human primates, also present logistical and ethical difficulties (5).

In this review, we evaluate the existing evidence for viral LAI prevention, including available vaccinations, post-exposure prophylaxis (PEP), and follow-up procedures following occupational exposure to Risk Groups 3 and 4 (RG3 and 4) viral infectious agents within human testing laboratories (risk group classification provided in Table 1). Specifically, we focus on high-consequence human viral pathogens associated with high fatality rates and/or severe neurological deficits. Where there is a discrepancy between risk group classifications, the highest risk group classification was chosen. As there is a paucity of evidence in human prophylactic studies, a summary of clinical and preclinical evidence is provided as an overview of PEP use in these uncommon yet significant exposure events, rather than providing recommendations. Decision-making should consider the level of evidence, the exposure event, and the specific clinical presentation.

TABLE 1.

Comparison of risk group designation for each virus by various guidelines^a

Virus	U.S. NIH (2024) (6)	Canada ePATHogen (7)	U.S. CDC select agent (8)	Australia/New Zealand 2243.3 (2022) (9)
Arenaviridae
Arenaviridae – Old World
Lassa virus	4	4	Yes	4
Lujo virus	–	–	Yes	–
Lymphocytic choriomeningitis virus (LCMV)	3 (neurotropic virus)	3 (neurotropic virus)	No	3 (neurotropic)
Arenaviridae – New World
Guanarito	4	4	Yes	4
Junin	4	4	Yes	4
Machupo	4	4	Yes	4
Sabia	4	4	Yes	4
Bunyaviricetes
CCHF	4	4	Yes	4
Hantaan and related viruses	3	3	No	3
Rift Valley Fever virus (Zinga virus)	3	–	Yes	3
Herpesviridae
Herpes virus simiae (B virus)	4	–	No	4
Filoviridae
Ebola virus	4	4	Yes (Tier 1)	4
Marburg virus	4	4	Yes (Tier 1)	4
Flaviviridae
Japanese encephalitis virus	3 (Japanese B encephalitis virus)	3 (Japanese B encephalitis virus)	No	3
Kyasanur Forest disease	4	4	Yes	4
Omsk hemorrhagic fever disease	4	4	Yes	4
St Louis encephalitis virus	3	3	No	3
Tick-borne encephalitis	4	4	Yes	4
West Nile virus	3	3	No	3
Yellow fever virus	3	3	No	3
Paramyxoviridae (Rhabdoviridae)
Rabies including fixed strain (CVS II)	2	3	No	3
ABLV	2	3	No	3
Hendra	4	4	Yes	4
Nipah	–	4	Yes (Tier 1)	4
Picornaviridae
Poliovirus	2	–	No	*b
Non-polio enteroviruses that cause encephalitis	2 (Echo virus and Coxsackie virus)	–	No	2

^a – indicates the distinction between Old World and New World Arenaviridae.

^b *, poliovirus is RG2 with additional biocontainment requirements as part of the global elimination program.

LAI INCIDENCE AND REPORTING

Incidence of LAIs with high-risk pathogens has been difficult to estimate due to lack of mandatory or standardized reporting. Historical surveys, including the largest survey to date conducted by Pike in 1976 (10), reported bacterial pathogens such as Brucella species as predominant causes of LAIs. Among RG3/4 pathogens, Venezuelan equine encephalitis virus was the commonest implicated viral LAI in this survey. The incidence and etiology of LAIs may differ regionally, with viruses, including the RG3 pathogen severe acute respiratory syndrome coronavirus (SARS-CoV), causing one-third of LAIs in the Asia-Pacific region in a review of published LAI reports between 1982 and 2016 (11). A more recent literature analysis of LAI between 2011 and 2020 was conducted to estimate the global number of published LAIs over a 10-year period, with 53 reports comprising 338 confirmed LAIs described (12). Of the 338 reported cases, 304 cases had an identifiable pathogen with the majority being bacterial. Of the documented viral LAIs, Crimean-Congo hemorrhagic fever virus (CCHFV) was the most commonly reported (25 cases).

The incidence of viral LAIs in high containment biologic laboratories is rare, with fewer than 250 cases reported over the last 35 years in a 2016 review (13). There have been suggestions that rates of LAIs have been decreasing, related to increased awareness of biohazards and improved biologic safety equipment and laboratory design (14). However, lack of standardized and accurate reporting of LAIs limits the ability to estimate the true incidence of these infections.

Efforts to make published reports of LAIs more accessible have led to the development of a comprehensive, searchable open-access database of published LAIs established by the American Biological Safety Association (ABSA), which captures peer-reviewed and published journal articles on LAIs (15). This database presents standardized reporting of LAIs, including descriptions of exposure incident and post-exposure prophylaxis provided. A search of “virus” in the database returned 190 published cases of viral LAI (16). This readily accessible and searchable database provides a useful resource for reviewing “lessons learned” from past exposure to laboratory pathogens in other centers. The Belgian Biosafety Server database of LAI and bio-incidents provides a summary of LAIs obtained from survey activities of laboratories in the Flemish region over 5-year intervals, as well as a global literature review of international reports (15, 17).

CURRENT LABORATORY SAFETY PRACTICES AND RISK MITIGATION

Modes of acquisition and risk mitigation

Exposure to laboratory pathogens can occur through various mechanisms, including inhalation of aerosolized particles, contact with the mucous membrane by splash, spills or touch, percutaneous self-inoculation (e.g., through needlestick injuries), direct contact with animal or insect vectors, or accidental ingestion. However, in published cases, more than 80% of reported LAI cases have had an unknown exposure route (12). The route of acquisition is important as the risk of infection and the manifestation of the infection can vary based on the mode; for example, mpox may manifest as a local lesion following direct contact with intact skin, versus systemic symptoms when the exposure involves deep inoculation of the skin (18). General preventative measures to ensure safe laboratory practices aim to address each potential mode of LAI acquisition. Where risk elimination is not possible, countermeasures to address the risks of handling pathogens that can lead to transmission should be incorporated into standard practice and safety procedures (19). These include targeted measures for specific routes of transmission such as operating within biosafety cabinets and use of emergency eyewash and decontamination stations to address the risk of droplet splashing and skin or mucus membrane exposure. The key elements of biosafety including appropriate facilities and equipment, personal protective equipment, safe operation rules, and management are essential (19). Appropriate biorisk and biosafety training must be provided to staff to ensure best practices and adherence to well-documented laboratory safety procedures (20).

A review conducted by Pedrosa and Cardoso assessed modes of acquisition relating to 219 viral LAIs published in 35 articles between 1930 and 2008 (21). Only cases where clear or probable laboratory exposure could be ascertained were included. The majority of LAIs related to arboviruses (including flaviviruses, alphaviruses, and bunyaviruses) were associated with aerosol exposure (84%), and the remainder were through percutaneous exposure. Aerosol-generating procedures included manipulation of viral suspensions of the infected animal tissue, working adjacent to virus culture, centrifugation accidents, and handling high-titer viral stocks (21). Inhalation was the main mode of acquisition in cases of lymphocytic choriomeningitis virus (LCMV), hantavirus, coxsackievirus, and SARS-CoV, with lack of appropriate droplet precautions cited in some of these cases.

Aerosolization of particles has occurred in the laboratory through accidents or malfunctions, e.g., centrifuge leak or spill, or dropping of sample plates or glassware (22). The use of class II Biosafety Cabinets (BSCs) with laminar flow is designed to protect the user as well as the environment and products from being contaminated with or exposed to potentially infectious aerosols (20). Nevertheless, laboratory errors or accidents can occur, which may lead to inadvertent release of infective aerosols, highlighting the need for additional precautionary measures such as individual PPE to minimize exposure risk. While there is a paucity of data reporting on LAI from specimen processing in the laboratory outside of high containment facilities, there are several reports of LAI from high-risk viral pathogens at the time of specimen collection, highlighting the risk inherent in samples with high-titer virus that may be handled at specimen collection or processing.

OVERVIEW OF APPROACH FOLLOWING EXPOSURE TO LABORATORY PATHOGENS

General first aid measures following exposure incidents

Following exposure, general first aid measures should be implemented and the laboratory staff member assessed by designated medical staff. It is important for clinicians to acknowledge the unique laboratory environment, including the possibility of acquisition through unconventional modes of transmission (e.g., aerosolization through centrifuge leakage), and potential high inoculum exposure that may shorten the incubation period (23). In general, baseline serology to the exposed pathogen should be taken to allow comparison with convalescent testing. PCR methods are usually available and are useful if symptoms develop or for follow-up testing.

Prompt first aid response is critical in reducing the risk of infection following potentially hazardous exposure to laboratory pathogens. For percutaneous injury, this includes appropriate wound or exposure site cleansing. Wounds and skin sites exposed to potentially infectious body fluids should be washed with soap and water for 15 minutes, in line with established procedures for occupational needlestick injuries (24). Although use is not contraindicated, the efficacy of antiseptics such as 2%–4% chlorhexidine or alcohol-based hand washes in preventing the transmission of blood-borne pathogens is unknown. Injection of antiseptics or disinfectants into the wound is not recommended. Expressing of fluid by squeezing the wound has also not been shown to reduce acquisition of blood-borne virus transmission (25). Mucus membrane exposure from splash or spills should be flushed thoroughly with water. Eyes should be irrigated with saline or water, and contact lenses should be removed.

The following section describes the testing and management of selected RG3 and RG4 viruses. An evidence-based summary of approaches used in different studies to testing and management following exposure to various RG3 and RG4 viruses is described in Tables 2 and 3, respectively.

TABLE 2.

Post-exposure management of RG3 organisms associated with encephalitis or severe neurological manifestations^c

Virus	Potential route of laboratory transmission	Licensed vaccines (pre-exposure)	Incubation period	Post-exposure prophylaxis	Initial and follow-up testinga	Established and investigational treatment for confirmed infection	References
Arenaviridae
Lymphocytic choriomeningitis virus (LCMV)	Percutaneous (viral culture, LCMV strain ARM53b variant clone 13, 106-7 PFU/mL); aerosols	None available	8–13 days	Not established	Baseline serology; serology (IFA and ELISA) IgM/G detected from 18 to 19 days post-exposure; IgM undetectable by 10 weeks; IgG persists at least 18 weeks; LCMV PCR (blood, CSF) if symptomatic	Supportive care; ribavirin [LOE: level 4, two case reports; one case reported IV 30 mg/kg, followed by 16 mg/kg QID for 4 days then 8 mg/kg TDS followed by oral 400 mg/600 mg]; favipiravir [LOE: level 5, animal study]	(23, 26–28)
Bunyaviricetes
Hantaan and related viruses	Percutaneous; aerosols (multiple reports of animal laboratory transmissions possibly attributed to aerosol or close contact)	Hantavax (used in Korea)	2 days to 2 months, most infections evident within 4 weeks	Ribavirin [LOE: two human cases; 400 mg tid for 20 days or 600 mg bid]	Serology at baseline and during follow-up (IFA, ELISA, and VNT); timing and duration of follow-up unclear; seropositivity in mild or asymptomatic laboratory workers has been described; PCR on blood and respiratory secretions if symptomatic	Ribavirin for HFRS – loading dose of 33 mg/ kg, followed by 16 mg/kg q6h on days 1–4, and then 8 mg/kg q8h on days 5–7 [LOE: level 2, double-blinded, randomized placebo-controlled trial: decreased mortality and improved outcomes in settings where dialysis was not available]; ribavirin is not recommended in HPS due to lack of evidence of benefit [LOE: level 3, two clinical trials which showed no mortality benefit]; favipiravir [LOE: level 5, animal studies]; ETAR [LOE: level 5, animal studies]	(29–40)
Rift Valley fever virus (Zinga virus)	Percutaneous; aerosols (laboratory manipulation of viral culture); mosquitos	No licensed human vaccines	2–6 days	Ribavirin [LOE: animal studies—suppressed viremia; loading dose of 50 mg/kg followed by 10 mg/kg at 8 h intervals for 9 days]; favipiravir [LOE: animal studies]	Serology at baseline and during follow-up (historical case of laboratory-acquired infection of unclear route, confirmed using lethal mice challenge study) RVF PCR on blood if symptoms develop	Supportive care	(41–47)
Flaviviridae
Yellow fever virus	Percutaneous; aerosols	Yellow fever live-attenuated vaccine (YFV-VAX) recommended for laboratory workers handling YFV-VAX	3–6 days	Not established	Baseline and follow up flavivirus serology in ~2–4 weeks (based on serodynamics of various flavivirus LAIs and natural infections. ELISA, IFA, VNT, and DEB-ELISA available, although the timing of seroconversion varies between antibody isotype); PCR and serological testing on blood, urine, CSF of symptomatic case	Supportive care; interferon [LOE: level 5, animal studies including non-human primate models]; sofosbuvir [LOE: level 5, animal studies]; ribavirin—not protective in non-human primates [LOE: level 5, animal models]	(48–55)
Japanese encephalitis virus	Percutaneous; aerosols or mosquito bite	JEV vaccination live-attenuated (Imojev) and inactivated vaccine (JEspect) recommended for laboratory workers handling JEV	4–15 days	Not established convalescent plasma not protective [LOE: double-blind placebo-RCT of 22 children, no significant difference in outcome]		Supportive care; no viral specific therapies in routine use	(11, 56–59)
West Nile virus	Percutaneous; aerosols; mucous membrane contact	None available	3–14 days	Not established		Supportive care; corticosteroids [LOE: level 4, case reports (18 patients)]	(60–66)
St. Louis encephalitis virus	Percutaneous; aerosols	None available	4–21 days	Not established		Supportive care	(21, 66, 67)
Paramyxoviridae (Rhabdoviridae)
Rabies including fixed strain (CVS II) ABLV	Bat bite; percutaneous; aerosols (processing of animal neuronal/brain tissue)	Rabies virus vaccination (Imovax)	Days to many years (most cases within 3 months)	Rabies immunoglobulin (20 IU/kg) [LOE: animal studies and human data of outcomes when not administered]; rabies vaccination [LOE: animal studies]; rabies monoclonal antibody (Rabishield and RabiMabs) under investigation [LOE: randomized phase III clinical trial]	Serology (VNT, RFFIT) at baseline to determine any pre-existing vaccine immunity; serology and PCR on CSF, saliva, brain tissue, and nape of neck biopsy of symptomatic cases; seroconversion in LAI can be late (weakly positive at 3 weeks)	Supportive care	(68–74)
Picornaviridae
Poliovirusb	Percutaneous; aerosols	Oral and inactivated polio vaccine. Booster inactivated polio vaccination recommended every 10 years	2–10 days	Not established	Daily testing (PCR and L20B viral culture) with stool specimens and throat swabs till culture negative on three consecutive samples (~7–8 days unless immunosuppressed); routine environmental (wastewater) surveillance may be considered in low vaccine coverage areas; baseline and follow-up serology (VNT) after 15–21 days	Supportive care and physical therapy; pocapavir (1600 mg daily for 14 days) [LOE: level 2, double-blind placebo-controlled study (n = 144) involving mOPV1 challenge; faster clearance with pocapavir (10 days) vs placebo (13 days)]; V-7404 (2,000 mg twice daily) [LOE: level 2, phase 1 clinical study (n = 72)]; anti-poliovirus antibody A12 [LOE: level 5, animal study]	(75–82)
Non-polio enteroviruses that cause encephalitis	Percutaneous; aerosols	EV-A71 vaccines licensed in China and Thailand Whole virus EV-A71 vaccines provide cross-genotype protection but not across other HFMD enteroviruses	2–10 days	Not established	PCR if symptoms develop—specimen guided by symptoms (usually nose/throat swab, vesicle swab, and fecal sample)	Supportive care; pocapavir [LOE: level 4, case reports]; corticosteroids [LOE: level 4, case series]; convalescent plasma [LOE: level 4, case series]; molnupiravir [LOE: level 5, animal study]; rupintrivir [LOE: level 5, animal study]	(10, 83–88)

^a Serological and PCR follow-up described in respective studies rather than recommendations.

^b Poliovirus is RG2 with additional biocontainment requirements as part of a global elimination program.

^c LOE, level of evidence; NT, viral neutralization test; DEB, ELISA defined epitope blocking ELISA; RFFIT, rapid fluorescent focus inhibition test; HFMD, hand foot and mouth disease; HFRS, hemorrhagic fever with renal syndrome; HPS, hantavirus pulmonary syndrome; ETAR, ethynyl-[1,2,4]triazole.

TABLE 3.

Post-exposure management of RG4 organisms associated with encephalitis or severe neurological manifestations^b

Virus	Potential route of laboratory transmission	Licensed vaccines (pre-exposure)	Incubation period	Post-exposure prophylaxis	Initial and follow-up testinga	Level of evidence for antivirals for confirmed infection	References
Arenaviridae
Lassa virus	Percutaneous; possible aerosol	None available	2–21 days	Oral ribavirin 35 mg/kg load (max 2.5 g), followed by 15 mg/kg (max 1 g) q8H for 10 days [LOE: level 4, case series]	RT-PCR testing on blood if symptoms develop; PCR testing of urine might increase sensitivity; baseline serology and after 2–3 weeks of exposure can be considered to diagnose asymptomatic infection	Ribavirin IV 30 mg/kg load (max 2 g), followed by 15 mg/kg (max 1 g) q6h for 4 days, and then 7.5 mg/kg (max 500 mg) q8h for 6 days [LOE: level 4, single prospective study, use of historic cohort as control group]; favipiravir with ribavirin [LOE: level 5, single case report of two patients]; favipiravir [LOE: level 5, animal studies]	(89–101)
Lujo virus	Percutaneous	None available	9–13 days	Not established	PCR testing on blood if symptoms develop	Supportive care; ribavirin [LOE: level 4, case series]	(102–104)
Sabia, Machupo, Junin, and Guanarito viruses	Percutaneous; aerosols	For Junin virus only – Candid 1 (licensed for use in Argentina)	5–21 days	Oral ribavirin [LOE: level 5, animal studies]; convalescent plasma from recovered patients in endemic areas for Machupo and Junin virus [LOE: level 4, case series]	PCR on blood if symptomatic; baseline and convalescent serology	Ribavirin [LOE: level 4; small prospective studies for Junin virus demonstrating potential benefit with clearance of viremia but no mortality benefit. Case reports for Sabia and Machupo virus; convalescent plasma for Junin virus [LOE: level 2, double-blind trial (n = 188) demonstrated mortality benefit if given within 8 days; treatment was associated with late neurologic complications]	(29, 89, 105–115)
Bunyaviricetes (Nairoviridae)
Crimean Congo hemorrhagic fever (CCHF) virus	Percutaneous; aerosols	None available	1–7 days	Oral ribavirin for 5–14 days (variable dose range; no consensus on dose or duration) [LOE: level 4, meta-analysis of case reports and case series (n = 175 in pooled analysis); 4/55 (7%) acquired infection in the ribavirin group vs 98/110 (89%) with no ribavirin]	RT-PCR on blood if symptoms develop; higher sensitivity over serology in early infection, particularly within 72 h symptom onset; serology–IgM peak at 2–3 weeks and may persist for several months	Ribavirin [LOE: level 3, a meta-analysis of RCTs and observational studies (n = 1287 in pooled analysis) reported 1.7-fold higher mortality in patients not treated with ribavirin; another meta-analysis (n = 748 in pooled analysis) reported significant uncertainty in efficacy due to critical risk of bias in available studies]; convalescent serum [LOE: level 4, case reports]; favipiravir [LOE: level 5, animal studies]	(116–119)
Flaviviridae
Tick-borne encephalitis virus (TBEV)	Percutaneous; aerosols	Inactivated TBEV vaccine (FSME-Immun, Encepur, TBE-Moscow, EnceVir)	4-28 days	TBEV-specific immunoglobulin used in Russia (not recommended elsewhere in Europe due to concerns regarding antibody- enhanced disease)	Baseline and follow-up serology, if vaccine not administered; confirmation by PCR on blood if symptomatic; serology on CSF if symptomatic	Supportive care; IVIg [LOE: level 5, single case report 400 mg/kg 5 days & animal studies]	(120–127)
Kyasanur Forest disease virus (KFDV)	Percutaneous; aerosols	Formalin-inactivated KFDV vaccine licensed in India	3–8 days	Not established	Baseline and follow-up serology; PCR on blood in the acute phase, up to 2 weeks	Supportive care	(120, 121)
Omsk hemorrhagic fever virus	Percutaneous; aerosols	None available	3–8 days	Not established	Baseline and follow-up serology (ELISA) PCR on blood	Supportive care	(122, 128)
Filoviridae
Ebola virus	Percutaneous	rVSV-ZEBOV (Ervebo) for Zaire Ebola virus	2–21 days	Monoclonal antibodies – ansuvimab (mAb114) and REGN-EB3 for Zaire Ebola virus [LOE: level 4, case report of 23 unvaccinated contacts of a confirmed case]; rVSV-ZEBOV used for PEP for Zaire Ebola virus [LOE: level 4, case report of 27 individuals]	Baseline and follow-up serology if vaccine not administered; RT-PCR for symptomatic patients; surveillance PCR has been performed for patients admitted to hospital following high-risk exposure	Monoclonal antibodies ansuvimab (mAb114) and REGN-EB3 for Zaire Ebola virus [LOE: level 2, phase 2-3 clinical trial (n = 681) demonstrated mortality benefit at 28 days]; investigational monoclonal antibodies (MBP-431) with or without remdesivir [LOE: level 5, protection demonstrated in primate models with Sudan Ebola virus]	(129–136)
Marburg virus	Percutaneous	None available	2–21 days	Investigational monoclonal antibody MR191-N [LOE: level 5, case report (n = 1 human case); protective in primate model]	Baseline and follow-up serology; PCR on blood if symptoms develop (oral swabs and tissue can also be tested)	Supportive care; monoclonal antibody MR191-N [LOE: level 5, animal studies including non-human primate models]	(137–139)
Herpesviridae
Herpes virus simiae (B virus)	Percutaneous; mucosal contact; aerosols	None available	5–21 days	Oral valaciclovir 1 gram TDS for 14 days [LOE: level 5, animal studies. PEP recommended by the U.S. CDC for high-risk exposures; topical cidofovir under investigation [LOE: level 5, animal studies]	Baseline and follow-up serology at 3–6 weeks, and 3 months (if antivirals administered); PCR testing on site of inoculation after cleansing; PCR from the wound, conjunctiva, and oropharynx 1–2 weeks after exposure or after antiviral discontinuation	Intravenous ganciclovir [LOE: level 4, case reports]	(140, 141)
Paramyxoviridae
Nipah virus	Percutaneous; aerosols	No licensed human vaccines	3–30 days	Human monoclonal, m102.4, used for both NiV and HeV exposures [LOE: level 4, case reports and primate models; phase 1 study demonstrating safety in humans]	PCR testing on blood and CSF if symptomatic; baseline and follow-up serology at 3 weeks (ELISA and virus neutralization assay)	Supportive care; ribavirin [LOE: level 3, open label trial (n = 140), lower mortality (32%) with ribavirin compared to control (54%)]; remdesivir [LOE: level 5, animal studies including non-human primate models]	(142–145)
Hendra virus (HeV)	Percutaneous; aerosols	No licensed human vaccines; Equivac administered to horses for prevention of HeV	2–14 days	Human monoclonal, m102.4 (see above); ribavirin [LOE: level 4, case report of 4 individuals exposed to HeV, three did not develop disease]	Baseline and follow-up serology at 21 and 42 days	Supportive care	(142–149)

^a Serological and PCR follow-up described in respective studies rather than recommendations.

^b LOE, level of evidence; IVIg, intravenous immunoglobulin; ELISA, enzyme-linked immunosorbent assay.

ARENAVIRIDAE—OLD WORLD

Viruses in the Arenaviridae family are grouped into Old World and New World, based on genetic and epidemiologic relationships (150). Old World arenaviruses are generally restricted to the African continent, with the exception of lymphocytic choriomeningitis virus (LCMV), which can be found worldwide.

Lymphocytic choriomeningitis virus

LCMV is a rodent-borne arenavirus that can cause human zoonotic infection through exposure to infected secretions from mice, rats, and hamsters, either through direct contact or aerosol spread. Infection with LCMV can result in asymptomatic infection (one-third of cases), non-specific self-limited febrile illness, or severe infection with aseptic meningitis or encephalitis (151). In immunocompetent individuals, infection is usually self-limiting with expected full recovery. The mortality rate has been reported to be <1% (151, 152). There have been several reports of LCMV infection in pregnancy, newborns, and in utero, which have been associated with severe adverse outcomes, including miscarriage, fetal death, or long-term neurological sequelae (153).

There are numerous published reports of LAI with LCMV, including several cases of LCMV causing meningitis following acquisition through needlestick injury (23, 26), direct contact with infected animal secretions, or possible aerosol spread (154). There are currently no available vaccinations for LCMV and no available clinical data on post-exposure prophylaxis.

Where available, testing should include serum collection for LCMV serology. The patient should be counseled regarding possible symptoms that may develop, noting the usual incubation period of 8–13 days (152). Follow-up after 1–3 weeks should be offered for repeat serology to assess for seroconversion (26, 27). A delay in seroconversion can occur in immunosuppressed patients, and a longer period of serological follow-up should be considered (151).

If symptoms develop, testing for LCMV with PCR should be performed. LCMV has been detected in blood by PCR in a case of aseptic meningitis following occupational exposure (23). PCR should also be performed on CSF if features of central nervous system involvement are present, although sensitivity appears to be lower and duration of PCR positivity in CSF may be short (151). In the event of a confirmed infection, treatment is usually supportive. Ribavirin has been used in cases of infection in immunocompromised solid organ transplant recipients (27) and in a case of LCMV LAI in an immunocompetent individual who developed visual impairment (26); however, its efficacy in these cases is unknown. Data to support use of ribavirin are based on in vitro studies demonstrating antiviral activity in the cell culture (155), as well as demonstrated efficacy in Lassa fever (156). For the same reasons, insufficient data currently exist to make recommendations for use of ribavirin for post-exposure prophylaxis, particularly in view of known ribavirin toxicity including hemolytic anemia.

Favipiravir is a newer antiviral agent targeting RNA-dependent RNA polymerase (RdRp) to inhibit viral replication and has been evaluated in clinical trials for use in influenza and SARS-CoV-2. Favipiravir has demonstrated higher potency compared to ribavirin in LCMV-infected mice (28); however, its use in humans for LCMV infection has not been evaluated. As such, favipiravir use for LCMV treatment is not currently recommended, pending further data in humans.

Lassa virus

Lassa virus (LASV) is the causative agent of Lassa fever and an arenavirus endemic to parts of West Africa associated with Mastomys rodents (156). It is considered an RG4 organism. Infection with LASV is usually asymptomatic or may lead to a mild febrile illness (80% of cases); however, severe cases may develop, leading to hemorrhagic shock (157). Sensorineural hearing loss is a common complication (158). The case fatality rate (CFR) has been estimated to be 1%–2% (156); however, CFR in hospitalized patients is significantly higher (15%–30%) (159), and a CFR of 35% has been reported for imported cases in nonendemic areas (160). The incubation period for Lassa fever ranges from 2 days to 3 weeks (157).

Besides an early report in 1970 of a laboratory worker who acquired Lassa fever while working with infected mice (161), there have been no recent published cases of LAI with LASV and no reports of LAI from high-containment biologic laboratories. There are currently no licensed vaccinations for Lassa fever; however, there are several vaccine candidates in preclinical development or undergoing Phase I clinical trials (162).

Given the high mortality risk, post-exposure prophylaxis with ribavirin has been recommended following significant exposure to Lassa virus (e.g., penetrating needlestick injury or mucous membrane exposure) with a proposed guideline for post-exposure prophylaxis published by Bausch et al. (89). However, data to support ribavirin use for PEP are limited to case series (cumulative 30 people from three published case series received ribavirin following high-risk LASV exposure did not develop Lassa fever) (90–92). A proposed dosing regimen is oral ribavirin 35 mg/kg loading dose (maximum dose, 2.5 g), followed by 15 mg/kg (maximum dose, 1 g) three times a day for 10 days (89). Dosing should ultimately be tailored based on the exposed individual’s specific factors such as weight, kidney function, and liver function and taking into account other potential drug interactions.

The preferred method of detection of LASV is through reverse-transcription-PCR (RT-PCR) in blood (93), which should be performed if symptoms develop. RT-PCR is preferred over serology due to increased sensitivity during the early stages of the disease (within 2 weeks after symptom onset) (93). Several RT-PCR assays are commercially available for detection of LASV RNA; however, only a few have obtained regulatory approval for clinical use (163). Persistence of RNA in bodily fluids, particularly urine and seminal fluid, for many months has been described (164). No formal recommendations exist on post-exposure monitoring. In reported cases of high-risk contact to LASV, active monitoring with twice-daily temperature monitoring for 21 days has been suggested, with RT-PCR used in symptomatic individuals (94, 105, 165). Serology at baseline and 2–3 weeks after exposure has also been suggested to diagnose asymptomatic infection (94, 165); however, lack of availability currently limits its use. Currently available serologic assays are research use only (RUO) and detect IgG/IgM to LASV glycoprotein (94).

In severe disease, intravenous ribavirin is recommended by the World Health Organization and the U.S. CDC (95, 96). These recommendations are largely based on a single prospective study that demonstrated mortality benefit in individuals receiving ribavirin who had an elevated aspartate aminotransferase (AST) level ≥150 IU/L at the time of hospital admission (CFR 5% [1/20] in the ribavirin group versus 55% [33/60] in the untreated group) (97). However, the recommendation for ribavirin use in Lassa fever has met recent controversy given the limitations of this single prospective study. These include non-randomization, use of a historic cohort as a control group, and incomplete or missing data. Additional results from this study were subsequently released, which identified similar benefit with ribavirin in the subgroup of individuals with AST ≥ 150 IU/L, but higher mortality when AST < 150 IU/L (98, 99). Other observational studies have suggested potential benefit with ribavirin, but the overall efficacy of ribavirin remains uncertain given critical risk of bias in these studies (98, 99).

Favipiravir has been used in the treatment of Lassa virus infection as part of combination therapy with ribavirin. In animal models, favipiravir has been tested alone and in combination with ribavirin in mice and showed favorable outcomes with survival of all treated mice and a rapid decline in viremia most evident with combination therapy (100). In macaques, high-dose favipiravir was associated with survival of all four treated macaques, while those that received low-dose treatment all succumbed to the infection (100). Favipiravir has been used in two human patients with PCR-confirmed Lassa virus infection, where it was administered on days 5 and 8 of infection for a short treatment course of 5 days. The treatment duration was limited by adverse effects of nausea, vomiting, and transaminitis, but both patients survived (101). Adverse reactions of these drugs are important considerations that may limit their use.

Lujo virus

Lujo virus is an Old World arenavirus similar to Lassa virus, which has caused small outbreaks in Lusaka, Zambia, and South Africa notably involving a cluster of healthcare workers in 2008, but no LAIs have been reported (102). The CFR from this cluster of five cases was 80%. Similar to Lassa virus, transmission is by direct contact with the blood and bodily fluids of infected patients. Rodents are suspected to be the primary source of animal spillover of this virus. The incubation period is 9–13 days. The symptomatology includes fever, headache, myalgia, pharyngitis, diarrhea, and rash. Hemorrhage and disseminated intravascular coagulopathy (DIC) have been described, which are not usually seen in Lassa virus infection (102).

Given the limited number of human cases, there is no established prophylaxis after exposure to Lujo virus. Diagnostic testing for symptomatic cases includes molecular diagnostic testing, particularly real-time PCR of blood in BSL-4 facilities after day 2 of illness (103, 104). There are no commercially available diagnostic assays. Ribavirin therapy has been used in the treatment of Lujo virus, with data limited to a case series of five patients, two received ribavirin, one of whom survived (102).

Mopeia virus

Mopeia virus (MOPV) is an arenavirus closely related to Lassa virus, but it is nonpathogenic in nonhuman primates, and natural infections in humans have not been reported (166, 167). MOPV can induce strong type I interferon responses in human endothelial cells and has been used commonly in the laboratory as a surrogate virus to study therapies and immune activation for Arenaviruses, particularly Lassa virus (168). Given that no clinical sequelae have been documented in humans, post-exposure prophylaxis or antiviral treatment is unlikely to be required in cases of laboratory exposure. MOPV is classified as an RG4 microorganism in Australia (169) and is designated RG3 by the CDC (20).

ARENAVIRIDAE—NEW WORLD

New World arenaviruses are maintained in rodent reservoir hosts found in North and South America (150). Several viruses in this group are known causes of hemorrhagic fever.

Sabia, Machupo, Junin, and Guanarito viruses

The highly virulent Sabia (SABV), Machupo (MACV), Junin (JUNV), and Guanarito (GTOV) viruses are causative agents of Brazilian, Bolivian, Argentinian, and Venezuelan hemorrhagic fevers, respectively. They are all considered RG4 organisms, and mortality has been reported to range from 15% to 50% (170). Infection is usually acquired through exposure to infected rodent excreta. Cases of LAI have been reported to SABV (106, 171), MACV (172), and a probable laboratory exposure to JUNV (173). The mechanism of spread in these cases was predominantly aerosol, including cases related to leakage of fluid during centrifugation of primary blood samples (172) or viral culture isolate material (106).

A live attenuated vaccination for JUNV (Candid 1) has demonstrated effectiveness in agricultural workers in Argentina (107) and has been part of the Argentine National Immunization Plan since 2007, offered primarily to rural populations in endemic areas (108). Candid 1 is not currently licensed in the United States, in part due to concerns about its attenuation resulting from only a single-nucleotide change, which carries potential risk for reversion to pathogenic state (174). There are currently no available vaccinations for SABV, MACV, or GTOV. Cross-protection from Candid 1 vaccination to other Arenaviridae is unknown.

The incubation period may range between 5 days and 3 weeks but may be shorter with high inoculum exposure (175). For cases of LAI, diagnosis has been made using reverse transcription PCR 3 days after symptom onset for SABV (106) or serology demonstrating seroconversion in paired acute and convalescent sera (106, 171, 172). There are no commercially available diagnostic assays for New World arenaviruses.

There are currently no clinical data on antiviral PEP following exposure to New World Arenaviridae. However, it has been suggested that ribavirin can be used following high-risk exposure (89, 109, 110), based on data for its use in Lassa fever (89), as well as in vitro and nonhuman primate models demonstrating the efficacy of ribavirin in early infection (111, 112). Adverse reactions to and poor tolerance of ribavirin may limit its use for PEP. There have also been case reports of successful use of immune convalescent plasma obtained from recovered patients in virus-endemic areas as PEP for MACV and JUNV (29); however, access to convalescent plasma may be challenging in non-endemic regions.

Use of ribavirin for confirmed JUNV infection has been evaluated in small prospective studies on advanced Argentinian hemorrhagic fever, demonstrating potential benefit with clearance of viremia in all cases treated with ribavirin but no observed reduction in mortality (113, 114). However, participants received treatment 8 days or more from symptom onset, and it is unknown if earlier treatment would result in improved outcomes. Case reports of ribavirin use have also been published for SABV (106) and MACV (115). Successful treatment of confirmed cases of Argentinian hemorrhagic fever has been demonstrated using immune convalescent plasma in a double-blind trial (n = 188) when initiated within 8 days; however, treatment was also associated with late neurological complications (176). The availability of immune convalescent plasma is likely to be limited, particularly in non-endemic areas and with reducing incidence from vaccination. Favipiravir has shown favorable activity against New World Arenaviridae in in vitro studies (177, 178) but has not yet been evaluated for clinical use.

BUNYAVIRICETES

The Bunyaviricetes class comprises several viral families including Arenaviridae (discussed above), Hantaviridae, Nairoviridae, and Phenuiviridae. Several viruses in this order have also been implicated in clinical viral hemorrhagic syndromes. These viruses were previously classified under the order Bunyavirales, prior to elevation of its status to Bunyaviricetes class in 2024 (179). Viruses in this class include Hantaviruses (Hantaviridae family), Crimean Congo Hemorrhagic Fever virus (CCHFV, Nairoviridae family), and Rift Valley fever virus (Phenuiviridae family).

Hantaviridae

Hantaviruses are a group of enveloped, negative-sense RNA viruses in the family Hantaviridae, genus Orthohantavirus. They are associated with two main clinical patterns: hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS) (180). The principal reservoirs for hantaviruses are rodents, including rats, mice, voles, shrews, and moles (30).

Species classification of Hantavirus can be made based on geographical location into Old World hantaviruses and New World hantaviruses and on the associated rodent family reservoir for the virus (30). Hantaviruses associated with the Muridae family and Arvicolinae subfamily of rodents are found in Europe and Asia and referred to as Old World Hantavirus species. Viruses that cause human infection associated with the Muridae family include Hantaan virus, Saaremaa virus, Dobrava-Belgrade virus, and Seoul virus. In the Arvicolinae group, Puumala virus and Tula virus cause human disease (30). Cricetidae, subfamily Neotominae/Sigmondtinae-associated Hantavirus species are found predominantly in the Americas and are referred to as New World Hantaviruses (181). Hantaviruses associated with Neotominae/Sigmodontinae rodents and which cause human infection include Andes, Bayou, Black Creek Canal, Laguna Negra, New York, Rio Mamore, and Sin Nombre viruses (181).

Old World Hantavirus usually causes HFRS, while New World Hantavirus causes predominantly HPS, with a shared underlying pathophysiology of increased vascular permeability of small vessels (30). HPS is associated with worse clinical outcomes and a CFR of 30% (182). It is characterized by a febrile prodrome followed by non-cardiogenic pulmonary edema with hypoxia and shock, and finally a period of diuresis. HFRS similarly has a febrile prodrome with myalgia, headache, blurred vision, and an erythematous rash, followed by hypotension, which may progress to shock and hemorrhage particularly in the kidneys. Diuresis usually marks the beginning of recovery in both HPS and HFRS (30). Transmission occurs from handling infected rodent secretions and excretions. The incubation period varies from 2 days to 2 months with most infections apparent 2–4 weeks post-exposure (30, 182).

Numerous cases of LAI from Hantavirus have been reported globally primarily in the setting of animal research laboratories housing rodents or handling rodent tissue (31–36). Inhalation of infectious aerosols, including from aerosol-generating procedures such as centrifugation and homogenization of tissue, is commonly reported (33, 34, 36). Diagnosis can be established through either serologic testing or detection of hantavirus RNA by RT-PCR (37). There are several serological methods for Hantavirus antibodies tests described, including immunofluorescence, hemagglutination-inhibition assays, enzyme-linked immunosorbent assays (ELISA), immunoblot, and complement fixation (30). Only neutralization assays can determine whether the causative hantavirus species as serologic cross-reactivity within the Orthohantavirus genus is common (183). IgM antibodies typically develop during the acute phase of illness and last 2–6 months, while IgG persists for years after infection (30). IgM false positivity has also been reported with commercial serologic assays when compared to a reference assay at the U.S. CDC (184). Nucleic acid detection by RT-PCR may also be performed on blood and lower respiratory tract samples in the viremic phase of acutely sick patients (30).

There is insufficient evidence for the routine use of ribavirin post-exposure prophylaxis, but it has been used in the USA in a laboratory exposure to New World Hantavirus with no clinical or laboratory evidence of infection following a 20-day course of post-exposure treatment (29). Other post-exposure therapeutic options such as neutralizing antibodies from convalescent plasma donations and murine monoclonal antibodies from immunized animals have been proposed; however, robust human safety data do not exist. Limited efficacy data in animal models (e.g., highly lethal model) suggest that some of the monoclonal antibodies may provide broad neutralization within the Orthohantavirus genus (38, 185, 186).

To prevent Hantavirus infection, vaccines against hantavirus are available in several parts of the world. Formalin- or b-propiolactone-inactivated monovalent or bivalent vaccines against Hantaan and/or Seoul virus have been used to immunize populations in high-endemicity regions in China and Korea (38). Hantavax is one such vaccine against Hantaan virus administered in Korea since 1990 (38). Clinical trial data support the need for multiple doses to achieve a neutralizing antibody response, with a three primary dose followed by a booster dose schedule offering the most immunogenic response when measured by neutralizing antibody assay (187). Other vaccines in development include DNA vaccines targeting the M gene expressing envelope glycoproteins and subunit vaccines targeting the N protein and virus-like proteins (38).

Specialized and high-level supportive care including expertise with ventilation, extracorporeal oxygenation, and renal replacement therapy is crucial for treatment of severe Hantavirus infection and has an impact on mortality outcomes (29, 30, 38). Evidence for the use of antiviral treatment such as ribavirin from patient reports is mixed but, in parts of Asia where HFRS occurs, ribavirin is frequently used (29, 38). Evidence for ribavirin therapy comes from a double-blinded randomized control trial for postexposure against Hantaan virus carried out in China where ribavirin was reported to decrease the mortality particularly when given within 4 days (39). Similarly, ribavirin given intravenously for 7 days for HFRS in Seoul in 34 patients with serologically confirmed infection showed milder disease and less complications compared with historic controls (40). Use against other Hantaviruses such as Puumala virus could not replicate this mortality benefit from ribavirin, and an increase in adverse effects was noted (188, 189). Furthermore, the timing of administration seems to be important with little benefit noted in the cardiopulmonary phase of HPS (30). Other antivirals such as favipiravir and ETAR have only been tested in animal models showing plausibility as a future therapeutic (38). Passive immunization through neutralizing antibodies has been effective in vitro and animal models, but human data are lacking, and availability of convalescent plasma may be limited (38). Other classes of therapeutics under investigation include murine monoclonal antibodies and small interfering RNAs, but currently, there is insufficient evidence for routine use (38).

Crimean-Congo hemorrhagic fever virus (CCHFV)

CCHF was first reported in agricultural workers in the Crimean Peninsula in 1944 and later in the Democratic Republic of Congo in 1969 (from a historical sample collected in 1956) but is now recognized to be endemic to many regions of the world including Africa, Eastern Europe, Russia, Middle East, and China (190–192). Transmission is most commonly from ixodid tick bites, and the virus is maintained in a tick-vertebrate-tick cycle with a range of wild and domestic animal hosts including cattle, sheep, goats, swine, and birds (193). Infected animals do not manifest disease symptoms (192). Transmission may also occur from directly handling infected animal tissue or blood during the viremic stage or consuming unpasteurized milk (190). Human-to-human transmission may occur from contact with blood, secretions, and horizontal transmission (192). The incubation period ranges from 1 to 5 days post-tick bite and 5–7 days following animal or infected human exposure (190, 192).

Clinical illness with CCHFV presents with an initial pre-hemorrhagic phase with a prodrome of sudden onset fever, photophobia, myalgia, headache, abdominal pain, gastrointestinal symptoms, and conjunctivitis lasting 4–5 days. The second phase, or hemorrhagic of the illness manifests with petechiae, ecchymosis, and hemorrhage from mucosa and internal organs may occur. Death in severely ill patients occurs from hypovolemic shock, DIC, alveolar hemorrhage, neurological complications, and multiorgan failure (190). The case fatality rate ranges widely from 3% to 40% (190, 192).

LAI from CCHFV, including fatal cases, has been reported and frequently relates to accidental inoculation from needlestick injury, handling blood samples without PPE and aerosolization of infectious particles from culture and high burden samples (29, 56, 128, 194). Consideration of ribavirin PEP is suggested in high-risk cases by the CDC, and literature on its use has reported significant reduction in disease progression in those who received ribavirin, although no randomized controlled study has been performed (29, 116, 194–197). A meta-analysis of case reports and case series including 175 healthcare workers exposed to CCHFV reported a significant reduction in development of CCHF in those who received ribavirin PEP (4 of 51 infected [7%]) compared to those who did not receive PEP (98 of 110 infected [89%]) (116). The duration of ribavirin therapy for post-exposure treatment varied from 5 to 14 days administered orally (29). In symptomatic patients, molecular testing by RT-PCR on blood is preferred due to improved sensitivity over serology in early stages of infection, particularly within the first 72 hours of symptom onset (117). Antibodies are detectable from about 7 days after onset of disease, with IgM peak 2–3 weeks after symptom onset, persisting for several months (117, 192, 193). Commercial serologic and RT-PCR assays are available for CCHFV; however, in-house developed assays are commonly used in reference laboratories (198).

There are no licensed vaccinations for CCHFV, but several vaccines are in preclinical development using different antigenic targets to elicit immune protection (199). Supportive care is important for management of CCHF and may include electrolyte balance, blood transfusion, and respiratory support (118). CCHF virus is susceptible to ribavirin and favipiravir in vitro and in animal models, but only ribavirin has been used clinically in human patients (29, 200, 201). The evidence for ribavirin use is largely based on case series with conflicting reports on the benefit of therapy. A meta-analysis of the efficacy of ribavirin found a decrease in mortality rate by 1.7-fold with the greatest benefit seen when ribavirin was initiated in the initial phase of illness during the pre-hemorrhagic stage (118). However, another meta-analysis reported significant uncertainty in ribavirin efficacy, given critical risk of bias due to confounding in the majority of non-randomized studies (202). Given the mixed data on efficacy, the WHO does not recommend ribavirin as standard of care therapy and suggests its use be done as part of a clinical trial, where possible (203). Benefits from convalescent serum from recovered patients containing anti-CCHF immunoglobulin have been reported in a limited number of cases and case series (119), though the limited availability of convalescent plasma is a significant barrier to large-scale adoption (201). Treatment with interferon therapy has shown in vitro antiviral activity (201); however, its use in a small case series in South Africa found no demonstrable benefit and was associated with severe adverse effects requiring interferon discontinuation (195). Other therapies under investigation include monoclonal antibodies, favipiravir, and 2′-deoxy-2′-fluorocytidine, with data currently limited to in vitro studies and animal studies (201), including nonhuman primate data demonstrating the antiviral activity of favipiravir against CCHFV (204).

Rift Valley fever virus

Rift Valley fever virus (RVFV) is an RNA virus transmitted to humans by mosquito bites from Aedes mosquitos (less commonly Anopheles and Culex species), as well as by broken skin direct inoculation or inhalation of aerosols when handling infected animal blood or tissue (205). Outbreaks in human and livestock have occurred in Africa and the Arabian Peninsula since identification of the virus in 1931 in Kenya (205). Humans are dead-end hosts, and disease manifests after 2–6 days of incubation as an influenza-like illness or asymptomatic infection in the majority of cases; retinitis, meningoencephalitis, or hemorrhagic fever are less common presentations. Human-to-human transmission has not been documented, but several cases of laboratory-acquired infection have been reported in the United States, including two fatal historical cases (41–43).

Prophylaxis against RVFV is not well established. Ribavirin has been shown to have efficacy in in vitro, but in vivo data from animal models suggest it may only delay disease onset (44). The antivirals favipiravir and benzavir-2 also show promise in pharmacokinetic and efficacy studies in mice and hamsters (45, 46). There are currently no licensed vaccines for RVFV, but formalin-inactivated vaccine TSI-GSD-200 and live-attenuated MP-12 vaccine have previously been evaluated in humans demonstrating an immune response to RVFV (206, 207). The most recent vaccine for RVFV under investigation is a simian adenovirus-vectored RVFV—ChAdOx1 RVF, which was found to be safe and immunogenic in a phase 1 human study (208).

Diagnosis of confirmed cases involves molecular detection and isolation of the virus and/or serological assays for detection of antibodies; however, the period of viremia with RVFV is short-lived after symptom onset. Serology for RVFV relies on immunofluorescence or ELISA methods for detection of IgM and/or IgG using recombinant proteins or whole inactivated virions. Serological tests, including detection of IgG seroconversion several weeks after an exposure, may play a role in confirming infection after a laboratory incident (47).

Treatment for RVFV is mainly supportive care with a number of therapeutics trialed including ribavirin, corticosteroids, convalescent plasma, phenytoin, and rifampicin without sufficient evidence to routinely recommend these therapies. Other antivirals, including favipiravir, are under investigation (209).

FILOVIRIDAE

Ebolavirus

Ebolavirus disease (EVD) results from infection with Ebola virus and has a reported case fatality rate of up to 50% (210). Major outbreaks of Ebola have predominantly involved Zaire ebolavirus (ZEBOV) and Sudan ebolavirus (SUDV). The incubation period ranges from 2 to 21 days with disease manifestations including fever, vomiting, diarrhea, rash, and bleeding (211).

Vaccination with rVSV-ZEBOV (Ervebo) has been approved by the U.S. Food and Drug Administration (FDA) as a single dose for protection against ZEBOV (129), and recommendations have been made by the U.S. CDC for vaccination of adults who are at the highest occupational risk of exposure to ZEBOV, including laboratory workers in BSL-4 facilities (212). The CDC has also endorsed booster dose administration of rVSV-ZEBOV for at-risk adults ≥ 6 months since prior vaccination (213). rVSV-ZEBOV has also been used as post-exposure prophylactic vaccination following cases of percutaneous exposure to ZEBOV in a BSL-4 laboratory worker in Germany (130) and in healthcare workers (131, 132).

Diagnosis of EVD relies on RT-PCR detection of Ebola virus in blood. PCR on oral swabs is insensitive and has limited diagnostic value (214). In cases of high-risk occupational exposure, daily surveillance PCR has been performed on inpatients for up to 10–14 days (133). Where available, serology should be performed at baseline and follow-up testing at 4–6 weeks to assess development of subclinical infection.

The monoclonal antibodies, ansuvimab (Mab114) and REGN-EB3, are FDA-approved and have demonstrated efficacy in treatment of confirmed EVD with ZEBOV (134). Both antibodies have been used as PEP in a prospective study of 23 non-vaccinated high-risk contacts who remained negative for ZEBOV during the follow-up period up to 14 days (135). Prior to the availability of these monoclonal antibodies, recombinant vesicular stomatitis virus-based ZEBOV (rVSV-ZEBOV) vaccination was used as PEP for several individuals following high-risk occupational exposure (133). There are currently no approved monoclonal antibodies or vaccinations available for SUDV or other Ebolavirus species. An investigational monoclonal antibody cocktail comprising two broadly neutralizing antibodies targeting Ebolavirus has demonstrated protection in rhesus macaques following lethal challenge with SUDV (136, 215). Protection was improved when co-administered with remdesivir (136). These have not yet been evaluated in human clinical trials.

Marburg virus

Marburg virus is in the genus Marburgvirus, and there are two species: Marburg virus and Ravn virus. Both are zoonotic viruses maintained in bats, primarily the Egyptian Rousettes (216). Human infection has only been reported in Africa or linked with exportation from Africa. Marburg virus was first reported in 1967 in West Germany linked with exportation of grivets (Old World monkeys) from Uganda for medical research as part of poliomyelitis vaccine development (217). Since 1967, fifteen other Marburg virus outbreaks have occurred. Exposure to deceased animals during autopsy resulted in an LAI from a breach in PPE and blood exposure. Since this time, 15 other outbreaks have occurred, including in South Africa (1975), Kenya (1980), the Democratic Republic of Congo (1998), Angola (2004), and most recently in 2023 involving Equatorial Guinea, Guinea, Ghana, and Tanzania (218, 219). LAIs apart from the 1967 German case include two other cases (one a fatal case) in Russian laboratories (216). A laboratory exposure in the United States where monoclonal antibody MR 191 N was administered has also been reported in the literature, with minimal additional information (137).

The clinical course of Marburg virus infection is similar to those with EVD, typically beginning with flu-like symptoms that progress to hemorrhagic manifestations (216). Laboratory findings include leukopenia with lymphocytopenia, thrombocytopenia, and prolonged coagulation times (220). Transaminitis and acute kidney injury have also been described as part of multiorgan involvement (216). The median incubation period is 7 days but ranges from 2 to a maximum of 26 days (216, 219). Infection is associated with a high fatality rate ranging from 20% to 90% based on outbreak data (219, 221).

Prevention of Marburg virus infections focuses on avoiding direct contact with oral secretions, urine, and fecal matter from infected bats, fruits contaminated by infected bats, infected animals, and bodily fluids from infected people. Candidate vaccines are being developed, including a modified adenovirus viral vector vaccine and modified vesicular stomatitis virus vector (222). Phase 1 human clinical studies have been conducted using a replication-deficient recombinant chimpanzee adenovirus type 2 vector vaccine (cAd4-Marburg) showing promising safety and immunogenicity with antibody response demonstrated at 4 weeks and persisting to 48 weeks (223).

Currently, there is no post-exposure prophylaxis treatment for Marburg virus that has been approved for humans, but there are a number of emerging therapeutic options being studied in primate models (220). Monoclonal antibodies designed to neutralize the virus, such as human anti-Marburgvirus GP monoclonal antibody, MR191-N, are provide protection in rhesus monkeys infected with the Angola strain of Marburg and have been used in one human case of laboratory exposure to Marburg virus, but the clinical details are not published (137). Small interfering RNAs are another therapy under investigation that work by inhibiting the expression of viral nucleoprotein (220). In primate models, small interfering RNAs, particularly those targeting the Marburg virus nucleoprotein, have been shown to be protective against the virus, particularly when administered in the first 3 days after exposure (220). Post-exposure vaccination with a viral vector such as recombinant vesicular stomatitis virus-based Marburg virus has also been trialed in a small number of primates with improved survival seen when administered hours after infection as benefit waned with time (220).

Laboratory confirmation of infection involves nucleic acid detection and antigen capture tests performed on blood samples; however, oral swabs and tissue may also be accepted. Serology for antibody detection may also be performed and is helpful in the convalescent phase of infection and retrospective diagnosis (138, 139). Treatment for Marburg virus focuses on supportive care as there are no standard-of-care treatments. Therapeutic options are similar to those described above for post-exposure prophylaxis and include monoclonal antibodies such as M 191 N and small integrated RNAs. Other treatments under investigation but lacking human data specific to Marburg virus include antivirals such as favipiravir, remdesivir, galidesivir, and interferon (137, 224).

FLAVIVIRIDAE

Kyasanur Forest virus

Kyasanur Forest virus (KFV) is transmitted by ticks and infected animal products and secretions particularly from small rodents, shrews, and monkeys. KFV causes a spectrum of illnesses from fever to hemorrhagic and neurological complications in humans usually 3–8 days after exposure (225). The disease is largely confined to India and, in particular, the Karnataka state, with seasonal peaks during the dry season (120).

LAI from KFV has been reported related to aerosolization during specimen or culture manipulation, direct inoculation from broken glassware, and needlestick injury (226–229). Vaccination as pre-exposure prophylaxis against KFV with a formalin-inactivated vaccine strain has only been utilized in India and is not licensed elsewhere. To achieve an immunogenic response, two doses of the primary regimen are required, as well as booster doses, as the duration of protection is short-lived (120). There is no other chemoprophylaxis described in the literature.

While there is guidance around BSL-4 handling of KFV and standard precautions when caring for patients and handling bodily secretions, the requirement for high containment in India where KFV circulates is difficult to meet due to limited resources. Risk minimization through immunization of staff and antibody monitoring as well as use of biosafety cabinet and PPE has been proposed to facilitate handling of specimens for human diagnosis in BSL2 facilities in India (230).

Diagnosis of infected patients can be made through RT-PCR on blood samples in the acute phase up to 2 weeks, and serology for detection of IgM and IgG by ELISA is most common, though hemagglutination inhibition, complement fixation, and viral neutralization assays are also described in the literature (120, 121).

Treatment for KFV is supportive with no established therapeutic options; however, sofosbuvir (an NS5 inhibitor) is under investigation (231).

Omsk hemorrhagic fever virus

Omsk hemorrhagic fever virus (OHFV) is part of the tick-borne encephalitis complex of flavivirus found in the Omsk region of Siberian Russia with a transmission cycle involving direct tick bites and contact with infected rodents and their secretions (232). Infection manifests as a febrile illness with myalgia, gastrointestinal symptoms, and epistaxis. Most patients recover, but one-third experience a complicated biphasic illness with encephalitis, nephritis, and/or pneumonia (122). The incubation period from exposure to onset is usually 3–8 days, and case fatality is low (232).

LAI from OHFV has been reported in Russia related to viral culture manipulation and likely aerosolization, but human-to-human cases have not been reported (128). Apart from precautionary measures taken when handling specimens in PC4 facilities, there are no specific prevention strategies or vaccinations. There is, however, animal data suggesting cross-neutralization of OHFV with inactivated whole virus tick-borne encephalitis vaccine (233).

Diagnosis of exposed individuals involves detection of OHFV RNA and serology assays looking for seroconversion or detection of OHFV-NS1 antigen or IgM/G antibodies (122). Treatment for OHFV is supportive only with no viral-specific treatments.

Tick-borne encephalitis virus

Tick-borne encephalitis virus (TBEV) is primarily transmitted by the bite of hard ticks within the Ixodidae family and less commonly via the consumption of unpasteurized animal milk products or contact with infected animal tissue. TBEV is found throughout Asia and Europe, with a seasonal predominance in warmer months (234). The incubation period from infection is 4–28 days, and the spectrum of illness ranges from asymptomatic to a nonspecific febrile illness or neuroinvasive disease with aseptic meningitis, encephalitis, or meningoencephalomyelitis. Outcomes of infection and case fatality rates vary depending on the subtype of TBEV (234).

LAI from TBEV has been described in unvaccinated laboratory personnel handling virus isolates—most often due to an aerosol generating procedure (235). Since the introduction of the inactivated TBEV vaccine, there has been a reduction in LAIs. The WHO position paper on tick-borne encephalitis recommends vaccination of all individuals living in highly endemic areas, targeted risk-based vaccination in moderate- to low-risk areas (236). Recommendations for immunization of laboratory workers vary by jurisdiction in Europe; vaccination is recommended by the US Advisory Committee on Immunization Practices for laboratory workers handling TBEV (123, 237). A three-dose primary vaccine course is recommended with a booster dose for those at ongoing risk 3 years after the primary course (124, 237). The serological correlate of protection is not known, so use of serology to guide the timing of booster vaccination is not advised (236).

The TBEV vaccine is not recommended for post-exposure prophylaxis due to the time course needed to induce an immune response (236). In Russia and Kazakhstan, TBEV-specific immunoglobulins are administered for post-exposure prophylaxis, but these are no longer used in Europe due to concerns regarding antibody-enhanced disease (125). Infection despite immunoglobulin therapy has been reported (126).

Following exposure, TBEV is diagnosed by PCR in the acute first phase of illness when the patient is viremic and by serology on serum and/or CSF looking for IgM and IgG seroconversion (127). PCR confirmation of TBEV in the tick may also be performed to confirm the exposure (125).

IVIg has been trialed for TBEV, but there is a lack of evidence to support routine administration of any immunomodulatory or antiviral therapy. The mainstay of treatment is supportive care (125, 127).

Yellow fever virus

Yellow fever virus (YFV) is transmitted by Aedes and Haemogogus mosquitos in tropical regions of Africa and South America: the transmission cycle usually involves nonhuman primates and mosquitos (rarely humans). Direct human-to-human infection is rare, but human-mosquito-human transmission occurs in outbreak settings (238). YFV causes a spectrum of illnesses from asymptomatic to fatal infection with multiorgan failure. The incubation period from exposure to infection is 3–6 days (239).

LAI from YFV was reported prior to available vaccines from handling biological samples from infected patients or animals. LAI from YFV is due to direct inoculation as well as inhalation of aerosols. The administration of live-attenuated YFV vaccination prophylaxis for laboratory workers is recommended by the WHO as well as regional and national bodies including the Pan American Health Organization and Australia National Health and Medical Research Council (NHMRC) for laboratory workers handling the virus, especially staff concentrating and culturing the virus (48–50). The recommended course is a single dose and repeat serology to check for neutralizing antibodies. Where serologic testing is not available, a booster dose every 10 years is recommended (240). Complications from vaccination—including hypersensitivity, neurotropic disease, and viscerotropic disease—are well described but uncommon and should be monitored (241). Apart from vaccination and vector control, no other pharmacotherapies are available for prophylaxis.

Following exposure, diagnosis is made by molecular testing for the virus as well as serology looking for the detection of IgM, IgG seroconversion, and/or a fourfold rise in antibody titers. Viral neutralization assays are the gold standard for serological diagnosis, but even with these assays, cross-reactivity with other flaviviruses (e.g., Powassan virus) can occur, as seen with other serology methodologies including hemagglutination, complement fixation, and ELISA testing, albeit much lesser when using neutralization assays (51). Immunohistochemistry on paraffin-embedded tissues and histopathology can also be performed to make a diagnosis.

Treatment for YFV focuses on supportive care. Agents with potential antiviral activity, including ribavirin, sofosbuvir, and interferon, have been studied in animal models, but there is insufficient evidence to support routine use (52–54).

Japanese encephalitis virus

Japanese encephalitis virus (JEV) is a neurotrophic flavivirus capable of causing acute encephalitis in infected individuals. Humans are a dead-end host for the virus. Other more common disease manifestations include asymptomatic infections or a febrile illness with headache (242). The virus is transmitted by Culex and Aedes mosquitos and usually circulates in a cycle involving waterbirds and pigs as amplifying hosts (243). Infections in humans occur from mosquito bites and (rarely) direct contact with infected animal tissues. The virus is endemic to Australia, Western Pacific Islands, and many areas of Asia including China and India (243). Disease patterns are linked with increased mosquito activity and warm weather, and hence the effects of climate change can impact geographical changes in the epidemiology of infection.

LAI from JEV has been reported relating to accidental percutaneous inoculation and inhalation (56). Vaccination is recommended for laboratory workers handling JEV, especially virus cultures and performing culture manipulation, as neutralizing antibodies have been demonstrated to protect animals against infection with a lethal challenge (57–59). There are different types of vaccines available, including live-attenuated (Imojev), live recombinant vaccine, inactivated mouse-brain-derived vaccine (JE-VAX), vero cell-culture-derived vaccine (Ixiaro), and inactivated vaccine (JEspect). There are differences in which vaccines are licensed and used in different settings, with JEspect and Imojev available in Australia (57). The immunization schedule for JEV vaccines also varies with the live-attenuated vaccine given as a single dose, while the inactivated vaccine requires a two-dose series with a booster given after 1 year if there is ongoing exposure risk (57).

Following an exposure, convalescent serum has been described for prophylaxis, but there is a lack of data outlining its efficacy, especially in vaccinated patients (244). There are currently no antivirals administered post-exposure. Exposed individuals should have acute and convalescent serology sent ~10 days after exposure noting that previous vaccination with JEV vaccines will impact the results (59, 245)

Diagnosis of symptomatic individuals in the early stage of infection involves RT-PCR testing on blood and urine as well as CSF if clinically indicated. Acute and convalescent serology on blood 14 days apart and CSF should also be performed (59). Serological cross-reactivity within the Japanese encephalitis serogroup (JEV, WNV, Usutu virus, and St. Louis encephalitis virus) is not uncommon, and accurate serological diagnosis of JEV is challenging and often requires co-assessment for other circulating flaviviruses within the same serogroup (51, 55). Many treatment options have been considered for JEV, including minocycline, IVIg, and interferon, but evidence to support their routine treatment is lacking (242, 246). Ribavirin has been found to be ineffective for treatment of JEV from clinical trials data (247).

West Nile virus (WNV)

Similar to JEV, WNV is a mosquito-borne flavivirus that circulates primarily between mosquitos and birds, which are amplifying hosts. Infected mosquitoes may also transmit the infection to humans and other mammals through biting. Humans and mammals are dead-end hosts (60). Infection in humans is most often asymptomatic but may manifest as a self-limiting febrile illness with headache, myalgia, arthralgia, and eye pain. In a minority of patients infected (~1%), neurological manifestations including meningitis, encephalitis, and acute flaccid paralysis may occur, which can be fatal (60). Transmission is most often through mosquito bites but can occur through exposure to infected animal tissues and secretions, blood transfusion, transplantation of infected organs, transplacental, and through breast milk (60). The incubation period from exposure to symptoms is 3–14 days.

LAI from WNV has been reported from accidental percutaneous inoculation, mucus membrane contact, and aerosol inhalation of culture isolates and infected tissues (61–64). The CDC has a guidance document on recommendations for protecting laboratory, field, and clinical workers from WNV exposure outlining appropriate use of PPE, sharp management, minimizing risk for aerosol-generating procedures, and use of mosquito repellents and protective clothing for field workers (248). There are no vaccinations in clinical use for prevention of WNV in humans, but there have been several candidate vaccines that have been assessed through clinical trials (249). Barriers to further development of these candidate vaccines have included safety concerns, cost, and demand given the unpredictability of WNV outbreaks (249). There is also no prophylaxis available after an exposure to WNV.

Management of an exposed individual should include immediate first aid as well as baseline serum specimen collection, which can be stored for later testing with a convalescent sample (65). Diagnosis of symptomatic individuals is made by virus isolation or PCR from blood and/or CSF. Seroconversion or the detection of WNV-specific IgM on serology may also be diagnostic with techniques including immunofluorescence, plaque reduction neutralization assay, and IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) (60, 66).

Treatment of WNV focuses on supportive care as there are no antiviral or immunomodulatory therapy routinely recommended. Clinical observations of improvement of neuroinvasive WNV with corticosteroid treatment have been noted across several reports in the literature, but the efficacy of corticosteroids needs to be evaluated in a controlled clinical trial to determine the true effect (250).

St. Louis Encephalitis virus

St. Louis Encephalitis virus (SLEV) is found in Northern, Southern, and Central America, with periodic epidemics associated with seasonal changes and the circulation of mosquitos in the warmer months transmitting the virus between the bird-mosquito cycles and rarely to humans who are dead-end hosts (251). Infected humans are almost always asymptomatic, but in ~1% of individuals, a febrile illness may develop or, rarely, acute neurological illness ranging from aseptic meningitis to encephalitis and meningoencephalitis (249).

LAIs from SLEV have been reported from percutaneous exposure and aerosols (21). There are no guidelines addressing the prevention and management of LAI from SLEV. There are no vaccines to prevent SLEV nor prophylactic treatment following exposure. Diagnosis of patients exposed to SLEV is made through serology testing on blood and CSF as well as direct detection by PCR and viral isolation from specimens taken in the acute phase (66). The period of incubation is 4 days to 3 weeks (67). Given this, a stored serum sample at the time of exposure should be considered for paired future testing. There are no targeted medications used for the treatment of SLEV in confirmed cases; supportive care is the mainstay of treatment.

HERPESVIRIDAE

Macacine alphaherpesvirus

Macacine alphaherpesvirus, also known as Monkey B virus, affects macaques but can be transmitted to humans by monkey bites and scratches or contact with body fluids and tissues of infected macaques. Infection in humans manifests with flu-like symptoms progressing to encephalitis, which is fatal in 80% of cases (140).

Infected macaques may not have any obvious signs of infection or may develop vesicular lesions in the mouth that eventually burst, scab, and heal without scarring. Along with the lesions, hepatic, renal, and central nervous system involvement can be observed (252). Monkey B virus infection is common in macaques (up to 97% seropositivity in captive populations), and seroprevalence increases with age (253).

LAIs have been described from percutaneous inoculation from bites and scratches in animal facilities, percutaneous inoculation from needlestick injury, cage scratches, and conjunctival and mucous membrane exposure from infected cell culture material, fluids, or autopsy material (141, 254, 255). Human-to-human infections are rare, with only one case reported from auto-inoculation of a skin cream shared between an infected case and spouse (256). The incubation period reported from exposure to infection is 5–21 days, with one case reported of latent infection and reactivation a decade after last documented exposure (141).

A management guideline for Monkey B virus has been published led by the B Virus Working Group convened by the Centers for Diseases Control and Prevention in 1999 (141). Preventative measures including appropriate PPE (including eyewear), staff education and training, and access to occupational health are emphasized. Post-exposure interventions, including immediate first-aid irrigation of the site for a minimum of 15 minutes with water or sterile saline if non-intact or mucosal site or with disinfectants including chlorhexidine and povidone-iodine for intact skin. Examination of the macaque for vesicular lesions and consideration of testing the primate by molecular and culture methods should be given if the exposure was a direct human-macaque incident. Serology on the primate is not useful due to the high incidence of infection in macaques, and negative serology does not rule out early infection (141).

Testing of the site of inoculation may be considered to help interpret the risk associated with the exposure, but this should not take precedence over wound first aid. A positive PCR or culture result from the wound does not prove infection. Serology testing at the time of the exposure and 3–6 weeks later should be performed (141). A competitive ELISA or Western blot is required to confirm positivity for Monkey B virus specifically and eliminate cross-reactivity with herpes simplex virus (141).

Post-exposure prophylaxis with oral valaciclovir 1 g three times a day for 2 weeks should be given within a few hours of exposure for patients where a breach to skin integrity has occurred, where there has been inadequate cleaning or mucosal exposure, where the site of inoculation is the head or neck, needlestick of deep puncture bite, any laceration where the macaque or exposure material has confirmed herpes B virus, or when the post-cleansing wound culture is positive for monkey herpes B virus (141). Topical cidofovir is under investigation for prophylactic administration at a wound site, but further data are required (140).

Following an exposure, vesicular lesions can occur at the site of inoculation for the infected case. As part of post-exposure follow-up, the wound should be monitored for the development of these lesions or symptoms of Monkey B virus. Serological follow-up should be extended to 3 months post-exposure if the 3–6 month serology is negative as antivirals can delay antibody production. Molecular testing on wound swabs, conjunctival, and oropharynx can also be considered at 1–2 weeks after antiviral discontinuation (141).

Treatment of cases suspected on clinical grounds with intravenous acyclovir is recommended. When there are neurological symptoms or a definitive diagnosis, IV ganciclovir is preferred (141).

PARAMYXOVIRIDAE

Nipah virus

Nipah virus (NiV) has been found in South and Southeast Asia, where it has been linked with outbreaks involving interaction between mammals (particularly pigs) and humans, as well as consumption of raw date palm sap contaminated by bat secretions (142, 257). Pteropus fruit bats are the reservoir host of NiV virus; mammals such as pigs and humans can be infected from contact with bats or their secretions. Other mammals such as dogs have also been reported as susceptible to NiV and capable of onward transmission to humans. Also, human-to-human transmission occurs from respiratory droplets and other bodily fluids (258). Infected humans may develop a febrile respiratory illness and, in severe cases, a fatal encephalitic illness (257, 259). The incubation time can vary from 3 to 30 days after exposure. Reactivation of latent infection months after exposure has also been reported (257).

Confirmed LAI from NiV has not been reported, but exposure events including mucus membrane contact, skin exposure to body fluids, and needlestick injury have been reported among healthcare workers (260). There are no guidelines specific to management of LAI, but a number of articles have outlined prevention measures for NiV, including animal and environmental control and use of PPE (259). With regard to prophylaxis post-exposure, a human monoclonal antibody named m102.4 targeting the ephrin receptor-binding sites of the G glycoprotein of NiV and Hendra virus (HeV), another Henipavirus, has been demonstrated in animal models of ferrets and monkeys to neutralize NiV after lethal challenge when administered soon after (143, 144). Human administration of m102.4 has been studied in a double-blinded placebo-controlled Phase 1 study finding the single and serial administration of m102.4 were safe and well tolerated (146). In Australia and the United States, the monoclonal antibody has been administered to a small number of patients on compassionate grounds for PEP following significant exposure to NiV or HeV (147). Vaccination for prevention is also under investigation, but currently, there is no approved human vaccine. Virus vector vaccines and soluble glycoprotein vaccines have been trialed in animal models and shown to be protective (144).

Following an exposure to NiV testing for infection is recommended, but precise guidance on which tests and what interval is lacking (142). Laboratory diagnosis of confirmed cases is made using direct virus detection by PCR and culture from CSF and blood and serology by ELISA and serum virus neutralization tests, which are the gold standard (142). Given this, it is reasonable to collect a serum sample for storage for later parallel serology testing. The timing of IgG positivity from outbreak studies was 100% of cases by day 18; 3 weeks may be an appropriate time frame for post-exposure follow-up testing (142).

Treatment of confirmed cases focuses primarily on supportive care. Apart from one open-label controlled study supporting the use of ribavirin to reduce mortality, there are no other antivirals in routine use (144, 145). Remdesivir has been evaluated in non-human primates challenged with a lethal dose of NiV and demonstrated complete protection (4/4 survival in the treated group vs 0/4 in untreated) (148). Efficacy of remdesivir against NiV in humans has not yet been evaluated.

Hendra virus

Pteropid fruit bats are the natural reservoir of Hendra virus (HeV) (261). The virus is transmitted to horses by contact or consumption of infected bat excretions, particularly urine. Infected horses may experience fatal encephalitis, and contact with infected horse tissues and fluids can result in human infection. Infected humans develop an influenza-like illness that can progress to respiratory distress or encephalitis, which can be a relapsing encephalitis (149). The incubation period from exposure to infection is reported to be a few days to 2 weeks (149). HeV has been found in Australia and linked with a number of equine cases and seven human cases with four deaths (261, 262).

Human infections from HeV have occurred mostly in veterinarians and animal workers in close contact with infected horses or their tissues post-mortem. No human-to-human transmission has been described. Management of a confirmed equine case is done by performing euthanasia to prevent onward transmission (149, 261, 263). The main prevention strategy in horses is the recombinant HeV G glycoprotein vaccine Equivac, which has been approved for use in Australia since 2015. It has high efficacy, with no confirmed cases of HeV in vaccinated equine (149).

Post-exposure management of cases is similar to NiV with human monoclonal antibody m102.4 administered to patients following significant exposure (146, 147). Ribavirin has also been administered following exposure to four cases, one of whom experienced a fatal encephalitis. More work is required to determine the efficacy of ribavirin (147). Australian guidelines on the management of exposed individuals advise a baseline serology sample and repeat at 21 and 42 days after exposure (147). There are no antiviral treatments used for confirmed HeV, and similar to NiV, the mainstay of treatment is supportive care.

PICORNAVIRIDAE

Poliovirus and neurotropic non-polio enteroviruses

Enteroviruses (EV) are a large group of small non-enveloped RNA viruses in the Picornaviridae family (264). Enteroviruses are responsible for a broad spectrum of clinical manifestations, which can affect almost all organ systems, including conjunctivitis, myocarditis, pneumonia, and hand, foot, and mouth disease (265). Among the most severe manifestations are neurological sequelae from neuroinvasion, which is characteristic of several strains, most notably polioviruses 1-3, EV-A71, EV-D68, EV-C105, and echovirus −7, 11, 19, 25, and 30 (264). Neurological manifestations include acute flaccid paralysis, aseptic meningitis, and encephalitis (266). Transmission of enterovirus is most commonly via the fecal-oral route or respiratory droplets. The incubation period ranges from 2 to 10 days (265, 267).

Poliovirus causes poliomyelitis, a painful paralytic myelitis that affects <1% of infected individuals (75). With the introduction of the polio vaccines, the oral live-attenuated vaccine and the inactivated polio vaccine, and the global efforts for polio eradication, wild-poliovirus (WPV) strain 2 and 3 have been eradicated, and only WPV-1 remains (268). The major challenge arising from the success of vaccinations has been the emergence of complications from the live-attenuated oral vaccine made with Sabin strains. Vaccine-derived poliovirus (VDPV) occurs in susceptible populations where the OPV is passed through multiple people over a prolonged period (269). Mutations of the virus occur, allowing neurovirulent virus to develop, particularly arising from the Sabin 2 strain (268). To tackle the rise of VDPV, several strategies have been tried, including removing Sabin virus 2, which is less genetically stable, and later introducing a novel oral polio vaccine containing a modified Sabin 2 strain with enhanced stability (269). Despite these efforts, VDPV remains a threat with ongoing potential for specimens containing these isolates to make their way into laboratories.

As part of containment measures for poliovirus, coordination, destruction, and control measures of laboratory isolates and specimens are required by the WHO Global Action Plan for Poliovirus Containment to limit the potential for accidental release of poliovirus (270). Poliovirus-Essential Facilities (PEF) with trained expert staff are stationed across the world with the key roles of facilitating polio confirmation and diagnosis, research, vaccine development, and clinical and environmental surveillance (270). LAI from poliovirus has been reported from direct inoculation of virus, aerosolization of culture isolates, and while handling infected animals where a cutaneous puncture occurred. These were more common in the pre-vaccine era but have been reported in vaccine production and research facilities (76, 77).

In addition to the routine course of polio vaccination, Australian guidelines recommend a booster dose of inactivated polio vaccination every 10 years for all laboratory workers in contact with specimens containing poliovirus (78). The polio eradication network outlines in the polio outbreak response plan the steps to be taken following an exposure to poliovirus dependent on the risk level. These strategies include isolation, management of feces, and enhanced cleaning. Also outlined is the suggested diagnostic surveillance for an exposed individual, which includes daily testing with stool specimens and throat swabs for 7 days and baseline serology to be repeated 15–21 days later (75, 79).

In confirmed infection, treatment focuses on supportive care and physical therapy. Antiviral therapies for poliovirus would be beneficial for treatment and post-exposure but are not yet standard of care. There are several antiviral agents that are being explored particularly for immunodeficiency-associated vaccine-derived poliovirus (iVDPV). iVDPV typically occurs in patients with B-cell immunodeficiency who, following vaccination with the oral polio virus, continue to excrete the vaccine virus for prolonged periods, increasing the risk of virulent mutations (80). Among the antiviral agents that have been evaluated is pocapavir, a capsid inhibitor that inhibits replication of the virus by preventing uncoating after virus attachment. Pocapavir has been evaluated in a randomized, double-blinded placebo-controlled human monovalent oral poliovirus vaccine type 1 (mOPV1) challenge where the test cohort received oral pocapavir or placebo after being immunized with an mOPV1, and time to clearance of virus from stool was measured. The pocapavir group cleared the virus more rapidly from the stool (median 10 days in the placebo group vs 13 days placebo), with half of the participants clearing the virus before the first placebo patient. There was an observation made of resistant virus emerging with therapy, which may have been due to transmission and reinfection within the clinical study unit (80). There is an ongoing phase II clinical trial of pocapavir in the treatment of poliomyelitis, with results much awaited (271). Due to concerns about resistance emerging, combination antiviral therapy has been proposed, with the leading candidate antiviral agent V-7404, which is a 3C protease inhibitor with potent in vitro activity and synergistic activity with pocapavir when tested against poliovirus (272, 273). A phase I study involving 72 participants showed that V-7404 was well tolerated and exhibited an acceptable safety and PK profile. The main complication to note was raised pancreatic enzyme levels without pancreatitis (81).

The other potential therapy under investigation are monoclonal antibodies against poliovirus such as anti-poliovirus antibody A12, a chimpanzee-human monoclonal, which has been evaluated in mice studies where it neutralized and protected mice against wild-type and VDPV strains (82). Similarly, human monoclonal antibody 9H2 also shows potential in in vitro studies with the ability to neutralize all three serotypes of poliovirus (274).

Neurotropic non-polio enteroviruses (NPEV) nosocomial outbreaks have been reported frequently, but reports of laboratory acquisition in the literature are scarce (10, 83). Within this collection of viruses, the only vaccine currently available is against EV-A71: there are three vaccines targeting EV-A71 available in China (84). These are alum-adjuvanted vaccine containing inactivated C4a sub-genotype, which have similar seroconversion rates. While the vaccine is effective against sub-genotypes of EV-A71, it does not provide protection more broadly against other genotype A enteroviruses (84, 275). Unlike poliovirus, there is no guidance on monitoring following an exposure event. Diagnosis of infection relies on molecular testing, mainly PCR, and less commonly viral culture. The optimal timing of diagnosis by molecular testing is usually early in the illness with the choice of specimen guided by the symptomatology and natural course of the virus. For many NPEV, this would include a nose/throat swab, vesicle swab, and fecal sample (85). Treatment of NPEV is most commonly supportive care (86, 265, 266). Adjunctive therapies such as corticosteroids, immunoglobulin, and convalescent plasma have been documented in the literature from case reports and series but lack high-quality evidence (86). Pleoconaril is one antiviral agent that has been evaluated in a clinical trial for EV-A71 neonatal sepsis, but it failed to show faster viral clearance and no survival benefit (276). Rupintrivir and molnupiravir have each been evaluated in animal studies and shown to be efficacious against EV-A71 (87, 88). Other treatments described include pocapavir for a patient with EV-D6, which was reported to have a favorable outcome when administered with immunoglobulin (277). Further research is needed to develop and evaluate antivirals for NPEV.

RHABDOVIRIDAE

Rabies virus and ABLV

Infection with rabies virus (RV) or Australian Bat Lyssa Virus (ABLV) can cause fatal encephalitis. The commonest method of transmission is through bites from an infected animal. Bites sustained to the head, neck, and upper body are associated with the highest risk of transmission. Other potential sources of acquisition include self-inoculation or skin breach from needles or cuts or mucus membrane exposure to contaminated tissue or fluids.

Aerosolization of RV is a significant risk in the laboratory setting, and cases of fatal LAI have been associated with aerosolization of the virus in pharmaceutical laboratories as part of work for vaccine development (68, 69). No fatal LAIs have been reported since the 1970 s despite reports of accidental exposures. This is likely due to a range of factors, including the availability of vaccinations, post-exposure care, and use of laboratory-adapted attenuated virus strains in diagnostics testing (69). Human-to-human RV transmission can occur from exposure to infected human fluids and tissues including saliva, urine, and nervous system tissue: appropriate facility containment (PC3) and PPE are essential. Transmission between humans has only been reported in the setting of organ transplantation and a suspected perinatal transmission (70). Other precautions to prevent LAI include handling specimens within a BSC , performing all aerosol generating manipulation within the biosafety cabinet including pipetting, tissue homogenization and opening of centrifuge gaskets, and substituting glassware with plastics (69). Disinfection of surfaces with quaternary ammonium, iodophors, 70% alcohol, and oxidizing agents, which inactivate rabies, is recommended.

The WHO guidance on rabies post-exposure treatment outlines three different categories (I–III) to stratify the risk of infection and management recommendations. Categories II and III involve a form of skin breach with increasing severity and risk and are associated with a need for first aid and post exposure therapy (70, 71).

Immediate first-aid following a laboratory exposure where there has been transdermal puncture or abrasion, exposure to broken skin or contamination of mucous membranes should include thorough wound cleansing with soap and water for 10–15 mins with care taken to avoid traumatizing the tissue further (69, 72). For wounds requiring suturing, this should be delayed where possible. Infiltration of the wound with human rabies immunoglobulin (RIG) (20 IU/kg) or horse immunoglobulin (400 IU/kg) is recommended in the WHO guidelines to neutralize RV at the site of skin breach (71, 73). Where there is no wound or if all possible RIG has not been injected into the wound site and the exposed individual has not been vaccinated, the remaining RIG should be administered intramuscularly (70). The benefit of administering RIG at a non-lesional site has been called into question especially given significant world-wide shortages of RIG and the effectiveness of other interventions such as rabies vaccination (72, 73). Human RIG is preferred since equine RIG is associated with side effects and hypersensitivity reactions.

Alternative post-exposure prophylaxis to passive immunization with RIG has been considered due to the challenges of sustaining a readily available supply especially in developing countries where rabies is endemic (74). In India, two monoclonal anti-rabies antibody treatments are available: Rabishield & RabiMabs. Rabishield has been studied in randomized trials of either administration of RIG or Rabishield monoclonals, and superior virus neutralizing activity was seen with the monoclonal antibody (74). The major concern is that the monoclonal only targets a single epitope compared with RIG, which has polyclonal binding to multiple epitopes to neutralize the virus. As such, Rabishield has already been found to be insufficient to neutralize some variant rabies viruses (74). Compared with Rabishield, RabiMab contains two mouse monoclonal antibodies targeting two distinct antigenic sites for broader application against variant and mutated virus and has been licensed in India since 2017. Phase 3 clinical trial data suggest it is non-inferior to RIG when given in combination with standard-of-care wound treatment and immunization (70, 74). Other novel monoclonals in development include recombinant monoclonals (74).

Vaccination forms an important part of both pre-exposure preparation and post-exposure management. The WHO recommends vaccination from inactivated RABV grown in cell culture or embryonated eggs. The virus obtained is concentrated, purified and inactivated, and characterized to ensure a potency of >/= 2.5 IU per dose. Laboratory workers, veterinarians, animal handlers, and other groups with a high occupational exposure to RV are recommended to be vaccinated. The route of administration can be either intramuscular or intradermal (ID) with evidence of efficacy from a lower dose and shorter course duration with intradermal vaccination. This has led to inclusion of the recommendation of a three-dose vaccine course given intradermally on days 0, 3, and 7 days or a 4 day intramuscular vaccination on days 0, 3, and 7 and between days 14 and 28 for post-exposure from a category III type exposure in the non-immune and a two-dose regimen when the patient has been immunized (70, 278). For pre-exposure vaccination in unimmunized at-risk individuals, a two-site ID vaccine on days 0 and 7 or IM vaccine on days 0 and 7 (70, 278) is recommended.

For individuals at ongoing risk of rabies exposure, serologic testing every 6–24 months is recommended to ensure adequate levels of neutralizing antibody protection; booster vaccination is offered if antibody titers fall below 0.5 IU/mL (69). If an immunized person is exposed, post-exposure vaccination is offered regardless of the antibody titer level to stimulate an amnestic antibody response (69, 72).

Following an exposure, the incubation period can vary from days to a year, with most cases symptomatic by 3 months (71). There are no tests for diagnosis before symptom onset. Diagnosis of symptomatic cases involves nucleic acid detection, viral isolation, direct antigen detection, and serology. Serology methods include ELISA as well as viral neutralization assays, including rapid fluorescent focus inhibition tests and fluorescent antibody virus neutralization, which is considered to be the gold standard neutralization test and is especially important for assessing vaccine efficacy (69). Ante-mortem biopsy of the nape of the neck and post-mortem brain biopsy for nucleic acid testing and fluorescent antibody testing are also helpful to establish the diagnosis (70).

Given the significant clinical similarities with RV, ABLV is managed and treated the same as rabies virus.

CONCLUSION

Staff working in BSL 3 and 4 facilities face unique biological hazards, which can pose a significant threat to human health. Knowledge of potential pathogens and preventative and post-exposure prophylactic measures is critical to occupational health and safety. When an exposure occurs, guidance on the therapeutic options, recommended follow-up, and clinical spectrum is essential for timely management. It is also important that with respect to local infectious diseases, physicians are aware of the work being undertaken in nearby high-containment laboratories and are involved in preparedness planning. Where possible, readily accessible stocks of relevant therapeutics and vaccines should be maintained to support rapid clinical response in the event of an exposure. This review provides an outline of the current evidence surrounding these high-risk viral pathogens, which may serve to guide institutions on implementing their own policies and procedures for occupational exposures relevant to their scope of practice. There is also a significant need for further research and development of antivirals, particularly broadly acting antivirals active across viral families. In addition, research protocols to establish pathways for rapid assessment of the clinical efficacy of antivirals in outbreak settings are needed. Both are essential for future pandemic preparedness, ensuring therapies for post-exposure prophylaxis and treatment of established infection are available, safe, and effective.

Biographies

Grace Butel-Simoes is undertaking dual training in Medical Microbiology and Infectious Diseases in Victoria, Australia after graduating from the University Of New South Wales, Sydney, Australia in 2015. This review was performed during her time training at the Victorian Infectious Diseases Reference Laboratory as the microbiology registrar. Her interests include microbial genomics and pathogenesis.

Michael A. Moso is an infectious diseases physician and clinician scientist based at The Royal Melbourne Hospital and The Peter Doherty Institute for Infection and Immunity in Melbourne, Australia. He received his medical degree at Monash University and completed specialist training in Infectious Diseases (FRACP, Australia) in 2023. He is currently completing his PhD at the University of Melbourne. Dr Moso has research interests in basic science and clinical virology, including HIV. His current research involves development of novel therapeutics using mRNA and lipid nanoparticle technology.

Prashanth Ramachandran is a neurologist specialising in neuroinfectious diseases and neuroimmunology. As a physician-scientist, he leads a laboratory investigating host-pathogen interactions and neuroimmune responses in central nervous system infections at the Peter Doherty Institute in Melbourne, Australia. He has published extensively in peer-reviewed journals and actively contributes to advancing the field through both basic science discoveries and translational research. His work aims to bridge the gap between laboratory discoveries and patient care.

Maryza Graham graduated with MBBS from The University of Melbourne in 1996. Maryza is currently a Medical Microbiologist at The Victorian Infectious Diseases Reference Laboratory and is also a Medical Microbiologist & Infectious Diseases Physician at Monash Health, Victoria’s largest health service since 2011. Previously Maryza was a Medical Microbiologist at the Microbiological Diagnostic Unit Public Health Laboratory. Maryza has been a member of the RCPA Microbiology Advisory Committee for 12 years and is currently Chair of the National RCPA Selective Reporting of Antimicrobials Guidelines Committee. Maryza has been a member of the ASA National EUCAST AST Committee for 6 years. Maryza is currently also a Member of the Melbourne Genomics Health Alliance and the RCPA Serology Advisory Committee. Maryza has a special interest in evaluating, optimising and publishing research on Microbiology laboratory process and result reporting for enhancing clinical impact.

Deborah A. Williamson is Dean of Medicine and Head of the School of Medicine at the University of St Andrews. A clinician-scientist with a strong background in microbiology and virology, she has held strategic leadership roles in academia and public health across Australia, New Zealand, and the UK. She has authored or co-authored over 300 publications, and her research focuses on applying innovative technologies, particularly genomics, to tackle infectious diseases.

Chuan Kok Lim is a medical virologist and Acting Director at the Victorian Infectious Diseases Reference Laboratory (VIDRL). Dr Lim is the Virology co-lead at the Peter Doherty Institute for Infections and Immunity and Co-Director for the Centre for Pathogen Genomics at the University of Melbourne. Dr Lim acquired his Bachelor of Medicine (MBBS, University of Adelaide) in 2004, Fellowship in Microbiology and Infectious Diseases (FRCPA and FRACP, Australia) in 2016, and PhD (University of Adelaide) in 2022 in basic virology and stem cell modelling. He supervises the Australian National High Security Quarantine Laboratory (NHSQL), a BSL4 facility at VIDRL. His research focuses on translational virology in public health, especially novel approaches in characterizing emerging pathogens with pandemic potentials. He leads a programme in developing viral genomics and innovative diagnostics to support outbreak response and public health policy-making.