Basics of Yellow Fever – i.e The Bare Minimum

Flaviviruses flabber my gaster. There are multiple of these that all look the same and all cause very similar diseases (read: hemorrhagic fever). Dengue, West Nile, and Zika are the more widely known given the recent outbreaks in the US of the latter 2. The one that is of historical (and epidemiological) importance is yellow fever. This is going to be a very basic overview of the disease. There are things to be said about diagnostics and vaccination, but this will be the topic of another post (I won’t talk about therapeutics, even the ones in study, as this is a changing topic).

Some Historical Context:

One of the prevalent theories is the virus originated from East and Central Africa and then made its way into the Americas via the slave trade (1, 2). Within the Eastern and Central parts of the continent, the virus made its way into Western Africa and then into the Carribean in the middle of the 18th century. During this time, sugar plantations that required large slave-labor population were established, and thus the slave trade brought in 12.5 million enslaved Africans across the Atlantic and into the Carribean.  The east african slave trade brought around the same number of enslaved Africans over twice the time span, with the direction being primarily trans-Saharan and across the Red Sea. Given that not many slaves were brought into eastern Asia and Oceania, it is likely this could account for its lack of presence in these places. One of the consequences is that most of the cases tend to arise from South America and Africa, and this can be seen in the genotypes of the virus. One of the unique things about Yellow Fever is that it tends to be fairly preserved (one of the reasons we are still able to use the vaccine 17D-YF vaccine to this day) and only 7 general genotypes are in circulation (it is more complicated than that, but it simplifies things a bit). For instance, an analysis of 79 yellow fever virus isolates collected from 1935-2001 in Brazil demonstrated that all but 2 isolates (one from 1983 and another one from 1975) formed one single monophyletic clade (3) while another study of 38 wild strains of YFV in Africa found 5 different genotypes, with nucleotide variations ranging from 8% to 25.8% (4):

Despite this being a disease localized to South America and Africa, there have been several outbreaks that have occurred in the United States, with at least 25 outbreaks in New York between 1668 and 1970 and a New Orleans outbreak in 1898 that caused around 4000 deaths (5). 

What Goes On:

There are 2 generally accepted cycles of disease: the jungle cycle and the urban cycle (5):

The jungle cycle (also called the sylvanic cycle) involves the propagation of the virus in primates while the urban cycle involves the propagation of the virus in humans. In order to propagate it, both mosquitoes and hosts need to have high levels of viremia for transmission to be effective. In South America, the majority of YF activity is found around Orinoco, Amazon, and Araguaia river basins.

What about the incubation period? This ranges from 3-6 days (6), with one study using aggregate historical data fitted into 4 models (7). Here, the extrinsic incubation (period that includes the intermediate host i.e. the vector) and the intrinsic incubation (period once in the definitive hosts) periods were calculated. The mean extrinsic incubation period ranged from 12-16 days, while that for the intrinsic period ranged from 4.3-5.6 days:

Notably, the extrinsic incubation period was significantly shorter as the temperature increased:

While we typically care more about the incubation period once inside the human, I think this illustrates the potential of this virus re-emerging during climate change. 

While there is no robust literature, several publications have reported several characteristics of the disease. There are 3 clinical periods: infection, remission, and intoxication. These are as follows (8, 9):

• Infection: high-level viremia, which leads into symptoms of fever, headache, malaise, weakness, lumbosacral pain (seriously, what’s with all these viruses causing back pain?), nausea and vomiting. This lasts for around 3 days. 
• Remission: you feel well for about 24hrs and up to 48hrs but then
• Intoxication: intoxicated with…cytokines! Jaundice, albuminuria, oliguria, cardiovascular instability, and hemorrhagic manifestation occur. This is when things get really bad and people tend to die by day 10. Liver damage happens at around this stage. This happens in 15% of cases and antibodies appear in the blood at this stage

A review (8) also noted several pathological manifestations, which include coagulative necrosis of hepatocytes, prolonged PT, PTT, and clotting times with reduction of clotting factors to <25% of normal, tubular necrosis with little glomerular changes, albuminuria/oliguria and azotemia, and degeneration of the AV conducting system. As you can see, this makes the diagnosis incredibly difficult to tease out as they can look like severe septic shock (multi-organ dysfunction syndrome), or other nice tropical infectious diseases such as malaria, leptospirosis, or other hemorrhagic fevers. 

Most Common Symptoms

In general, these tend to be non-specific. There are some trigger symptoms that someone may actually be on their way of having severe disease. In one series (10) of 72 patients with yellow fever, most patients presented with fever, myalgia, headache, abdominal pain, and jaundice. The mortality rate was 29.2%. In another study of 79 patients presenting to an infectious-disease ICU found that the most common signs and symptoms were fever, nausea, abdominal pain, and myalgias (11):

Another retrospective study of 251 cases with a case mortality of 44% had similar findings (12):

Risk Factors for Severe Disease:

I think this is an interesting topic, though in the end, it doesn’t even matter since the labs and clinical picture tend to guide you. Nevertheless, it is helpful to know who is going to go down the metaphorical cliff. In one Sao Paulo retrospective study of 72 cases (10), a higher proportion of patients who died had a total bilirubin >2mg/dL:

And, not surprisingly, those who died had lower platelets, higher transaminases, higher INR and creatinine:

Factors associated with death included age, AST and creatinine based on 2 models, with the first considering these factors as continuous variables while the second considered them as categorical values (i.e. they used cut-off points):

In another cohort of 79 ICU patients (11) found that 61% of patients having an AST >7000 IU/L, and 76% having some sort of renal dysfunction. 24% also had seizures, while 58% had serum lipase concentration three times above the ULN. 65% of these patients had hemorrhagic manifestations, most often GI bleeds and bleeding from venipuncture sites:

In another study (12), patients with moderate and severe disease had worse liver function test and creatinine, though notably those with moderate disease had worse disease than those with severe disease:

Risk factors in multivariate analysis for death in admitted patients included jaundice OR 19.55, 95% CI 4.26 – 89.65) and AST >1200 IU/l (OR 2.57, 95% CI 1.14-5.77). In a cohort study of 95 patients with suspected yellow fever found that age, neutrophilia, AST and yellow fever RNA viral load were associated with increased mortality (13): 

An event associated algorithm that included a cutoff of neutrophilia of >4000 cells/mL and viral load of 5.1 log10 copies/mL found these 2 values had a high sensitivity for predicting mortality in this cohort:

In general, it seems that any bilirubin higher than 2, severe transaminitis (in the thousands), rising BUN, and viremia tend to be associated with death. So there is that. No clear scoring system, but if you see someone with bad symptoms and these labs = badness.

Diagnosis and Viral Kinetics.

The diagnosis is actually quite difficult to make. Testing is limited to national or regional reference laboratories, making it difficult to get testing done in a timely matter. For one, Yellow Fever assays cross-react with other flaviviruses (looking at you, Dengue). For another, the isolation of viruses is actually quite difficult, with viral culture being labor-intensive and requiring biosafety precautions. Further, while the viral genome is quite preserved, not all assays catch the 7 genotypes  and there may be some  cross-reactivity with the 17D strain that is used for vaccination. PCR assay has been used more frequently, with four real-time PCR assays being studied and able to detect all strains.

During the early phase of infection, viremia tends to predominate, with IgM increasing at around day 5 (14). Some data suggest that RNA can be detected in the urine for the first 20 days. After this, anti-YFV IgM antibodies develop and can be detected up to 3 months, sometimes even up to 3 years in some cases. For instance, in a study of 40 patients from Atlanta, GA, 73% of those who got the 17D-YF vaccine had positive IgM antibodies 3-4 years after vaccination (15). Those who were positive after the time skip were more likely to have earlier detectable viremia, as well as a higher median peak viremia (2253 copies/mL vs 298 copies/mL). Moreover, virus specific neutralizing antibodies were higher in the IgM positive group:

Serology tends to be the mainstay of diagnosis, though a single IgG or IgM is not enough. A 4-fold rise in titers between acute and convalescent samples is usually required. IIF and ELISA are the most common techniques available. There are 4 commercially available IgM- or IgG assays, with high sensitivities in one assay (94.4% -94.7%). The plaque reduction neutralization (PRNT) assay, which detects neutralizing antibodies against a specific cell culture by the prevention of plaque formation, is the most specific but again, cross-reacts with other assays (6)

Another method of diagnosis is molecular testing, which is gaining favor over serologies. PCR is the most common method, with some that detect all 7 genotypes being available (14):

Newer types of PCR have also been explored. A study evaluating RT-LAMP (RT loop-mediated isothermal amplification) in 120 hamsters with samples obtained from South American countries found that RT-dLAMP (using degenerate primers i.e. non-specific primers) and RT-sLAMP (specific primers) had higher sensitivities compared to RT-PCR, RT-Hemi Nested-PCR, and qRT-PCR (16):

Of course, this was done in hamsters and lacked any of the African viruses, so it is difficult to make much of this. Given the state of the data,  the WHO recommends that RT-PCR be prioritized and IgM Elisa be done if the PCR is negative (17). IgM positivity should be treated as a presumptive infection, with a follow up convalescent sample drawn one week later and with differentiated IgM against other flaviviruses. 

References:

1. Cathey JT, Marr JS. Yellow fever, Asia and the East African slave trade. Trans R Soc Trop Med Hyg. 2014 May;108(5):252-7. doi: 10.1093/trstmh/tru043. Erratum in: Trans R Soc Trop Med Hyg. 2014 Aug;108(8):519. PMID: 24743951
2. Bennett, Raphael Dolin, Martin J. Blaser. Mandell, Douglas, And Bennett’s Principles and Practice of Infectious Diseases. Philadelphia, PA :Elsevier/Saunders, 2015.
3. Vasconcelos PF, Bryant JE, da Rosa TP, Tesh RB, Rodrigues SG, Barrett AD. Genetic divergence and dispersal of yellow fever virus, Brazil. Emerg Infect Dis. 2004 Sep;10(9):1578-84. doi: 10.3201/eid1009.040197. PMID: 15498159; PMCID: PMC3320275.
4. Mutebi JP, Wang H, Li L, Bryant JE, Barrett AD. Phylogenetic and evolutionary relationships among yellow fever virus isolates in Africa. J Virol. 2001 Aug;75(15):6999-7008. doi: 10.1128/JVI.75.15.6999-7008.2001. PMID: 11435580; PMCID: PMC114428.
5. Barrett AD, Higgs S. Yellow fever: a disease that has yet to be conquered. Annu Rev Entomol. 2007;52:209-29. doi: 10.1146/annurev.ento.52.110405.091454. PMID: 16913829.
6. Domingo C, Charrel RN, Schmidt-Chanasit J, Zeller H, Reusken C. Yellow fever in the diagnostics laboratory. Emerg Microbes Infect. 2018;7(1):129. Published 2018 Jul 12. doi:10.1038/s41426-018-0128-8
7. Johansson MA, Arana-Vizcarrondo N, Biggerstaff BJ, Staples JE. Incubation periods of Yellow fever virus. Am J Trop Med Hyg. 2010 Jul;83(1):183-8. doi: 10.4269/ajtmh.2010.09-0782. PMID: 20595499; PMCID: PMC2912597.
8. Monath TP. Yellow fever: a medically neglected disease. Report on a seminar. Rev Infect Dis. 1987 Jan-Feb;9(1):165-75. doi: 10.1093/clinids/9.1.165. PMID: 3547569.
9. Domingo C, Charrel RN, Schmidt-Chanasit J, Zeller H, Reusken C. Yellow fever in the diagnostics laboratory. Emerg Microbes Infect. 2018;7(1):129. Published 2018 Jul 12. doi:10.1038/s41426-018-0128-8
10. Ribeiro AF, Cavalin RF, Abdul Hamid Suleiman JM, Alves da Costa J, Januaria de Vasconcelos M, Sant’Ana Málaque CM, Sztajnbok J. Yellow Fever: Factors Associated with Death in a Hospital of Reference in Infectious Diseases, São Paulo, Brazil, 2018. Am J Trop Med Hyg. 2019 Jul;101(1):180-188. doi: 10.4269/ajtmh.18-0882. PMID: 31134884; PMCID: PMC6609182.
11. Ho YL, Joelsons D, Leite GFC, Malbouisson LMS, Song ATW, Perondi B, Andrade LC, Pinto LF, D’Albuquerque LAC, Segurado AAC; Hospital das Clínicas Yellow Fever Assistance Group. Severe yellow fever in Brazil: clinical characteristics and management. J Travel Med. 2019 Jun 11;26(5):taz040. doi: 10.1093/jtm/taz040. PMID: 31150098
12. Tuboi SH, Costa ZG, da Costa Vasconcelos PF, Hatch D. Clinical and epidemiological characteristics of yellow fever in Brazil: analysis of reported cases 1998-2002. Trans R Soc Trop Med Hyg. 2007 Feb;101(2):169-75. doi: 10.1016/j.trstmh.2006.04.001. Epub 2006 Jun 30. PMID: 16814821.
13. Kallas EG, D’Elia Zanella LGFAB, Moreira CHV, Buccheri R, Diniz GBF, Castiñeiras ACP, Costa PR, Dias JZC, Marmorato MP, Song ATW, Maestri A, Borges IC, Joelsons D, Cerqueira NB, Santiago E Souza NC, Morales Claro I, Sabino EC, Levi JE, Avelino-Silva VI, Ho YL. Predictors of mortality in patients with yellow fever: an observational cohort study. Lancet Infect Dis. 2019 Jul;19(7):750-758. doi: 10.1016/S1473-3099(19)30125-2. Epub 2019 May 16. Erratum in: Lancet Infect Dis. 2019 Nov;19(11):e370. PMID: 31104909.
14. Waggoner JJ, Rojas A, Pinsky BA. Yellow Fever Virus: Diagnostics for a Persistent Arboviral Threat. J Clin Microbiol. 2018 Sep 25;56(10):e00827-18. doi: 10.1128/JCM.00827-18. PMID: 30021822; PMCID: PMC6156298.
15. Gibney KB, Edupuganti S, Panella AJ, Kosoy OI, Delorey MJ, Lanciotti RS, Mulligan MJ, Fischer M, Staples JE. Detection of anti-yellow fever virus immunoglobulin m antibodies at 3-4 years following yellow fever vaccination. Am J Trop Med Hyg. 2012 Dec;87(6):1112-5. doi: 10.4269/ajtmh.2012.12-0182. Epub 2012 Oct 29. PMID: 23109371; PMCID: PMC3516084.
16. Nunes MR, Vianez JL Jr, Nunes KN, da Silva SP, Lima CP, Guzman H, Martins LC, Carvalho VL, Tesh RB, Vasconcelos PF. Analysis of a Reverse Transcription Loop-mediated Isothermal Amplification (RT-LAMP) for yellow fever diagnostic. J Virol Methods. 2015 Dec 15;226:40-51. doi: 10.1016/j.jviromet.2015.10.003. Epub 2015 Oct 13. PMID: 26459206.