《1. Introduction》

1. Introduction

In general, identifying the first infected case (patient zero) is key in tracing the origin of a virus; however, doing so is extremely challenging. Despite extensive efforts, scientists have not yet identified patient zero for the 1918 influenza pandemic, human immunodeficiency virus (HIV), or H1N1 influenza in 2009, and patient zero for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is likely to remain unidentified as well. The challenge in identifying the origin of SARS-CoV-2 is that a great deal of interdisciplinary research is required; in particular, if patient zero was asymptomatic or had very mild symptoms, he or she may not have seen a doctor or generated a medical record. As a result, patient zero could forever remain unidentified. Therefore, what roadmap could be followed to skip over patient zero while still recognizing the origin of the virus?

Here, we propose a new virus transmission route (Fig. 1) by focusing on environmental media containing viruses such as SARS-CoV-2 or RaTG3-related bat-borne coronavirus (Bat-CoV), hereafter termed as the "environmental quasi-host.” We propose reasons why the environmental quasi-host is likely to be a key node in helping recognize the origin of SARS-CoV-2.

《Fig. 1》

Fig. 1. The SARS-CoV-2 transmission pathway.

Viral transmission to humans occurs via natural host–human contact or environmental quasi-host–human contact, where the environmental quasi-host might be water, soil, or food contaminated by an animal host’s urine, saliva, feces, or secretions. Many researchers believe that SARS-CoV-2 may have come from the wild animal market. Nevertheless, they have focused on the natural host–human pathway [1–3], while ignoring the natural host–environmental quasi-host–human pathway.

Is it possible that SARS-CoV-2 infected patient zero through contact with an environmental quasi-host? With rapid industrialization and globalization, contacts between humans and environmental quasi-hosts are greatly increasing, as human activity spaces strongly overlap with animals’ living spaces. Moreover, viruses can survive for a long time in certain environmental media [4–6]. Many viral outbreaks in humans have been caused by direct human contact with environmental media containing a virus, such as virus-carrying water and soil, rather than by direct contact with a natural host [7–10].

Based on the following pieces of evidence from recent research and other viral transmission pathways, we consider that SARSCoV-2 could have been transmitted from an environmental quasi-host.

《2. SARS-CoV-2 detection in various environmental media》

2. SARS-CoV-2 detection in various environmental media

SARS-CoV-2 has been detected in various environmental media (Table 1 [11–22]), including wastewater, soil, floor surfaces, door handles, sinks, lockers, tables, windows, and packages, to name just a few. Between February and March of 2020, Liu and colleagues [11] at Wuhan University in China demonstrated the presence of SARS-CoV-2 RNA in the air by setting up aerosol capture devices in and around two hospitals. Ong’s group [12] detected SARS-CoV-2 on environmental surfaces in patients’ rooms and toilets. SARS-CoV-2 has also been detected in wastewater at Schiphol Airport in Tilburg, the Netherlands [13]. SARS-CoV-2 may exist in the habitats of species that are natural hosts for SARS-CoV-2. Therefore, further examination of environmental media in natural habitats for SARS-CoV-2 is needed.

《Table 1》

Table 1 SARS-CoV-2 detected in environmental media [11–22].

《3. Long-term virus survival in environmental media》

3. Long-term virus survival in environmental media

Viruses can survive in environmental media for hundreds or even thousands of days and remain infectious under suitable conditions, which are often reported to be low temperatures, relatively closed conditions, less disturbed conditions, and highly heterogeneous environmental media. Mollivirus sibericum, which has been preserved in permafrost for 30 000 years, is still capable of infection after resuscitation [23]. Porcine parvovirus can survive in soil for more than 43 weeks [6], and poliovirus remains stable and active at 1 °C for 75 days [24]. In groundwater, human norovirus still has 10% activity after 1266 days [25]. In mineral water, hepatitis A virus and poliovirus only have a small reduction in infectivity for one year at 4 °C [4]. In contaminated water, norovirus can still be detected after 1343 days [5].

We have analyzed 482 scholarly papers published in the past 120 years (Table 2 [26–122]), which study the survival time of 116 different strains of viruses. From a statistical perspective, over 84% of the 116 different strains of viruses can survive for more than one week (Fig. 2 [26–122]). With the rapid development of global transportation, viruses in environmental media can be carried from one place in the world to another in days or weeks; thus, the origin of a virus could be far away from the location of its breakout. As the phylogenetic characteristics of a virus may greatly affect its survival time in environment media, the phylogenetic characteristics of viruses require further study.

《Table 2》

Table 2 Virus survival times in environmental media [26–122].

《Fig. 2》

Fig. 2. The distribution of the survival times of the 116 studied viruses [26–122].

Existing studies have confirmed that SARS-CoV-2 is likely to exist for a long time in septic tanks and other soil-containing solid media, as well as in the ground [22]. The Singapore National Center for Infectious Diseases and the Defense Science Organization (DSO) National Laboratories have detected the virus in the residence rooms of COVID-2019 patients; floor surfaces had the highest positive viral signal, exceeding those of toilets, door handles, sinks, lockers, tables, and windows [12]. SARS-CoV-2 was found to remain viable in aerosols throughout the experiment (3 h), with a reduction in infectious titer from 103.5 to 102.7 median tissue culture infective dose (TCID50) per liter of air [77]. Based on these findings, SARS-CoV-2 may exist and survive for a long time in habitat and activity place of wildlife, especially in places with low temperatures and low levels of light.

《4. Viral outbreaks in humans caused by direct contact with environmental media rather than contact with a natural host》

4. Viral outbreaks in humans caused by direct contact with environmental media rather than contact with a natural host

By analyzing the literature published in the past 120 years, we found at least 198 viral infection cases with 28 different strains of viruses that occurred through direct contact with environmental media (Table 3 [123–318]). Some of these cases were statistically derived from data in order to obtain a correlation between environmental media and viral transmission, and many were derived from investigations of environmental media that recognized the route or host of viral transmission. For example:

(1) A 44-year-old woman from Colorado, USA, suffered from Marburg disease in 2008 after returning home from a two-week tour in Uganda. This disease is caused by a virus that belongs to the same family as the Ebola virus, one of the deadliest pathogens to humans. Scientists sequenced the gene of an Egyptian fruit bat in a cave in Uganda and believed that she was infected by the virus when she touched a rock covered with bat feces while visiting the python cave [8–10].

(2) The transmission route of the Ebola virus has been confirmed as the human consumption of fruit contaminated by fruit bat feces [7].

(3) No less than five infectious disease incidents have occurred in China since 2009 due to drinking groundwater containing a virus that ended up affecting thousands of people. For example, an outbreak of gastroenteritis occurred in Hebei, China, in the winter of 2014–2015. The nucleotide sequence of the norovirus extracted from clinical and water samples had 99% homology with the strain of Beijing/CHN/2015, which confirmed that the outbreak was waterborne. This is an excellent example of finding the route of virus transmission by investigating environmental media [154,176,194,244,319].

《Table 3》

Table 3 Cases of virus infection caused by direct human contact with environmental media [123–318].

 

(4) Airborne transmission is an important mode of virus transmission, and at least six different cases of viruses infecting humans through airborne transmission have been reported. Alsved and colleagues took air samples from the surrounding environment of patients with norovirus infection and analyzed the norovirus RNA in the samples by reverse transcription polymerase chain reaction (RT-PCR). They detected norovirus RNA in some air samples, suggesting that air pollution from vomiting is an important source of norovirus [170,265,273,276,292,295].

Insights from a statistical perspective provide evidence for linkages between the environment and epidemics:

(1) Eight out of the 11 first reported human cases of Ebola occurred in areas with high levels of forest destruction, where the forests were the habitats of bats carrying the Ebola virus [320].

(2) The migration trajectory of ticks in damaged forest areas is significantly correlated to the distribution and morbidity of Kyasanur forest disease [321] and Lyme disease [322]. Moreover, habitat destruction increases both the survival pressure of wild animals and the viral load of urine and saliva secretions [323].

《5. Viruses in many animals might transmit to humans through multiple pathways》

5. Viruses in many animals might transmit to humans through multiple pathways

The order Nidovirales, sub-family Orthocoronavirinae, family Coronaviridae is composed of four genera: α-coronavirus, β-coronavirus, γ-coronavirus, and δ-coronavirus. SARS-CoV-2 belongs to the subgenus Sarbecovirus of the genus β-coronavirus, to which SARS-CoV and MERS-CoV also belong. Coronaviruses (CoVs) infect humans as well as domestic and wild animal species, with infections remaining sub-clinical in most cases [324–326]. α-coronavirus and β-coronavirus usually infect mammals, with a probable origin of bats, while γ-coronavirus and δ-coronavirus mainly infect birds, and sometimes mammals, and might originate from swine [327– 329]. A long list of animal species has been reported as intermediate hosts, such as dogs, cats, cattle, horses, camels, rodents, rabbits, pangolins, mink, snakes, frogs, marmots, hedgehogs, and ferrets [324,330–336]. Thus, there could be multiple viral transmission pathways from different animals to humans. The three outbreak points of coronavirus in China—namely, the livestock markets in Guangdong in 2003, the Huanan Seafood Market in Wuhan at the end of 2019, and the Xinfadi Seafood Market in Beijing in June 2020—are all related to animal markets. Civet cats and camels have been demonstrated to transmit SARS-CoV or MERS-CoV to humans, which provides an important hint of virus transmission directly from animals. However, it remains unclear which animal could be the main intermediate host of SARS-CoV-2, although positive viral RNA signals were detected in seafood markets and on the chopping boards of salmon. In 1983, Lidgerding and Hetrick [337] first reported the replication of a coronavirus in a fish cell line. Furthermore, Sano et al.[338] successfully isolated a coronavirus from common carp (Cyprinus carpio) in 1988, which induced hepatic, renal, and intestinal necrosis in experimentally infected fish. Miyazaki et al. [339] found a corona-like virus in color carp (Cyprinus carpio) in 2000, which caused dermal lesion and necrosis in internal organs.

Based on the aforementioned pieces of evidence, we propose that an environmental quasi-host can infect a human, and that there are two transmission routes of SARS-CoV-2:

(1) Natural hosts (animals with the virus)–environmental quasi-host (animal feces/water, soil and food contaminated by animals’ urine, saliva, feces, and secretions)–patient zero (infected or virus-carrying human who came into contact with the environmental quasi-host while traveling or working in the wild)–back to home or human habitations–outbreak of COVID-19.

(2) Natural host (animals with the virus)–environmental quasihost (fruit, food, or meat contaminated by animals’ urine, saliva, feces, and secretions)–transported to different regions or countries– patient zero (infected or virus-carrying human who came into contact with or ate the environmental quasi-host)–outbreak of COVID-19.

To summarize, it is imperative to investigate environmental quasi-hosts in order to source track the origin of SARS-CoV-2 through our two suggested transmission routes. Given the need to trace the virus around the world to prevent further pandemics, global collaboration is required not only to identify the origin of the virus, but also to fundamentally protect the existence and development of species. Doing so will proactively conserve and restore habitats for species, and serve as a key strategy for preempting the next pandemic.

《Acknowledgements》

Acknowledgements

We acknowledge the fund by Chinese Academy of Engineering (2020-ZD-15) for financial support of this work.

《Compliance with ethics guidelines》

Compliance with ethics guidelines

Miao Li, Yunfeng Yang, Yun Lu, Dayi Zhang, Yi Liu, Xiaofeng Cui, Lei Yang, Ruiping Liu, Jianguo Liu, Guanghe Li, and Jiuhui Qu declare that they have no conflict of interest or financial conflicts to disclose.