September 1, 2010

—Liz Highleyman
HCV Advocate

The hepatitis C virus (HCV) is relatively hardy compared with some other viruses, which has implications for transmission and prevention. HCV may survive outside the body for days in dried blood on surfaces, or for months in a liquid medium under favorable conditions. Paradoxically, however, HCV has proven difficult to maintain in laboratory cell cultures, which has hampered research on potential treatments.

Survival in the Lab

Due to the difficulty of maintaining viable HCV in the laboratory, researchers have relied on “replicon” models of a specific strain of HCV (JFH-1 genotype 2a) in a liver cancer cell line. But this year researchers from Massachusetts Institute of Technology and Rockefeller University finally found a way to sustain viral replication in healthy liver cells for up to three weeks.

As described in the February 16, 2010 Proceedings of the National Academy of Sciences, Sangeeta Bhatia and colleagues developed a way to maintain healthy hepatocytes, or liver cells, for four to six weeks by precisely arranging them on a specially patterned plate and mixing them with fibroblast cells that support their growth. The liver cells could then be infected with HCV for two to three weeks, giving time to study response to candidate drugs. The HCV strain used for this research came from a Japanese patient with fulminant hepatitis; the researchers hope to modify their system to maintain a genotype 1 HCV strain, the most common type in the U.S. and the hardest to treat.

Survival on Surfaces

According to the U.S. Centers for Disease Control and Prevention (CDC), HCV can survive on environmental surfaces at room temperature for at least 16 hours but no longer than four days. The more fragile HIV virus, in contrast, only lives on surfaces for a few hours, while influenza viruses may survive for several hours up to about a day.

The CDC estimate is based on a study by Kris Krawczynski and colleagues, presented at the 2003 American Society for the Study of Liver Diseases (AASLD) meeting and published in the May 2007 issue of Infection Control and Hospital Epidemiology. The researchers examined the stability of genotype 1a HCV in dried blood plasma from an infected chimpanzee.

Plasma samples were dried in test tubes overnight (for about 16 hours) then either rehydrated immediately using sterile water and stored at -70ºC (about -160ºF, the temperature of biomedical research freezers), or put in a controlled environment chamber with 42% humidity at 25ºC (77ºF, room temperature) for four or seven days before rehydration. The rehydrated virus was then injected into a different chimp to see whether it remained infectious.

HCV RNA (genetic material) was detectable in plasma dried overnight and stored for seven days, though viral load decreased by 1 log, or ten-fold, compared with the original plasma sample. The test chimpanzee injected with virus dried overnight developed detectable HCV RNA and elevated alanine aminotransferase (ALT), became HCV antibody positive, and had detectable HCV antigen in liver cells. Injection of rehydrated virus stored for four or seven days, however, did not lead to infection. The researchers therefore concluded that HCV could remain infectious on surfaces outside the body somewhere between 16 hours and four days.

Viability outside the body, however, can vary widely depending on conditions. Viruses survive longer on hard surfaces such as stainless steel and less time on soft surfaces like fabric. HCV can live longer at cooler temperatures and prefers humidity to dry conditions.

Survival in Liquid

HCV survives longer in liquids than it does when dried on surfaces. In one recent study, described in the June 15, 2010 Journal of Infectious Diseases, Sandra Ciesek from Hannover Medical School in Germany and colleagues looked at the environmental stability and infectivity of HCV grown in a laboratory cell culture, as well as its susceptibility to chemical disinfectants. The researchers measured changes in viral load and introduced recovered HCV RNA into cultured Huh7.5 liver cancer cells to test for infectiousness.

In a liquid environment, HCV was detectable for up to five months at lower temperatures. However, the researchers noted that the risk of HCV infection may not accurately be reflected by measuring HCV RNA levels, because viral infectivity and viral load were not directly correlated. Further, they found that various alcohols and commercially available antiseptics reduced HCV to undetectable levels, though diluting hand disinfectants reduced their virucidal activity.

In another study published in the February 2010 Virology Journal, Hongshuo Song from Peking University and colleges found that JFH-1 cell culture-derived HCV could survive in liquid culture medium for two days at 37ºC (98ºF, body temperature) and 16 days at 25ºC, but was relatively stable at 4ºC (about 40º, average refrigerator temperature) without major loss of infectivity for at least six weeks.

This cell culture-derived HCV was vulnerable to heat; infectious virus could be inactivated in four minutes at 65ºC (about 150ºF) or eight minutes at 60ºC (140ºF), but this took 40 minutes at 56ºC (about 130ºF). Ultraviolet light efficiently inactivated HCV within two minutes. Exposures to formaldehyde and various detergents destroyed infectious HCV effectively in both culture medium and human serum.

HCV’s ability to live for a prolonged period in liquid blood underlies its transmission via nasal drug use (HCV RNA was detected on 5% of straws from hepatitis C patients who “snorted air”—simulating drug use—in one recent study), tattooing, sharing personal care equipment such as razors, childbirth, certain sexual activities, and re-use of medical equipment in healthcare settings.

Epidemiologic studies show that hepatitis C prevalence is higher among people who have undergone various medical procedures including kidney dialysis, indicating that HCV can spread from one patient to another via contaminated equipment if proper infection control practices are not followed. In 2008, for example, several patients at a Las Vegas endoscopy clinic contracted hepatitis C when clinicians gave multiple people injections from the same vials of anesthesia medication.

Survival in Syringes

HCV survival in blood in syringes is a key concern, given that sharing needles for drug injection is the most common route of hepatitis C transmission. In a presentation at the 17th Conference on Retroviruses and Opportunistic Infections in February, Elijah Paintsil from Yale University School of Medicine reported findings from a laboratory study looking at how long HCV can live in syringes.

The researchers first filled syringes with HCV-infected blood and depressed the plunger, simulating what happens when a user “boots,” or draws blood up into a syringe to mix with drugs and then reinjects it. Either immediately or after storing for up to two months at various temperatures, the team flushed out the syringes and attempted to grow recovered virus in genotype 2 HCV in cell cultures. They analyzed both low-volume (2 microliter) insulin syringes with permanently attached needles and high-volume (32 microliter) tuberculin syringes with detachable needles.

In the low-volume syringes, the likelihood of finding infectious HCV declined rapidly, with no viable virus recovered after one day of storage at 37ºC or three days at 22ºC (72ºF). At 4ºC, viable virus could be detected in two-thirds of syringes after one day of storage, about 25% after three days, and about 5% after seven days.

But in high-volume syringes, infectious HCV could still be recovered from nearly all syringes stored at 4ºC for seven days, from about half of those stored for 35 days, and from about 10% even after 63 days. At higher temperatures of 22ºC or 37ºC, viable HCV could still be recovered from a small percentage of syringes after two months.

The longer survival of HCV in syringes helps explain why HCV transmission occurs ten times more often than HIV transmission from accidental needle sticks, and why harm reduction measures such as needle exchange have reduced HIV incidence more than new HCV incidence.

At an accompanying press conference Paintsil said that while it might be advisable for needle exchange programs to offer smaller insulin syringes, some individuals (for example, transgender people who inject hormones) want larger syringes, and the most important thing is to provide enough so that people never have to share.

Understanding how long HCV can survive outside the body can inform practices to reduce the risk of viral transmission. According to Krawczynski and colleagues, “The potential for HCV to survive in the environment re-emphasizes the importance of cleaning and disinfection procedures, safe therapeutic injection practices, and harm reduction counseling and services for injection drug users.”



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