CLINICAL MICROBIOLOGY REVIEWS, Apr. 1990, p. 120-131 Vol. 3, No. 2 0893-8512/90/020120-12$02.00/0 Copyright C) 1990, American Society for Microbiology Transport of Viral Specimens F. BRENT JOHNSON Department of Microbiology, Brigham Young University, Provo, Utah 84602 INTRODUCTION .......................................................................... 120 TRANSPORT SYSTEMS.......................................................................... 120 Swabs ........................................................................... 121 Swab-Tube Combinations ........................................................................... 121 Liquid Media .......................................................................... 122 Cell culture medium .......................................................................... 122 BSS and charcoal .......................................................................... 122 Broth-based media ........................................................................... 124 Bentonite-containing media .......................................................................... 124 Buffered sorbitol and sucrose-based solutions ........................................................................ 124 Transporters .......................................................................... 125 Viruses Adsorbed onto Filters .......................................................................... 125 Virus Stability in Transport Media: a Comparison .................................................................... 126 CONDITIONS FOR SHIPPING OR TRANSPORT OF SPECIMENS TO THE LABORATORY ........... 127 Temperature and Time.......................................................................... 127 Regulations .......................................................................... 127 PRESERVATION AND STORAGE OF VIRUSES ....................................................................... 127 RECOMMENDATIONS AND FUTURE DIRECTIONS ................................................................ 129 CONCLUSIONS .......................................................................... 129 LITERATURE CITED.......................................................................... 129 INTRODUCTION isolation in the laboratory. In some situations, the virus can be transported in the specimen itself. Examples of this Viral agents vary widely in composition, structure, mor- include urine, cerebral spinal fluid, blood, washings of the phology, size, and stability. For those viruses that infect respiratory tract, and stool (16). A variety of viruses can be humans, the loosely packaged enveloped viruses are gener- isolated from urine during phases of an infection when they ally more labile than the nonenveloped dense viruses. How- are being shed from the urinary system. Similarly, viruses ever, some enveloped viruses, such as herpes simplex vi- are found in cerebral spinal fluid, blood, and stool during ruses (HSV), are surprisingly stable in the nonfrozen state certain phases of some infections. These kinds of specimens when placed in a suitable holding environment. It is com- can be transported without placing them in an exogenous monly recommended that clinical specimens for virus isola- transport medium. tion either be inoculated into cell culture at bedside or be The purpose of this article is to review the transport media immediately placed in a refrigerator or on wet ice and then that have been reported in the literature and have been used inoculated into cell culture with as little delay as possible. successfully to carry infectious viruses from the patient to many common agents withstand However, in reality viral the laboratory for culture. both storage at room temperature and transport for extended periods of time (days) when placed in a suitable transport medium. Perhaps better and more complete diagnostic analysis of TRANSPORT SYSTEMS some infectious diseases would be accomplished if it were more widely recognized that viral specimens can be col- lected and transported to the laboratory for successful The constituents of suitable viral transport media are testing. The infectivity of viruses does, in fact, decrease over designed to provide an isotonic solution containing protec- time and generally the decay rate is a function of tempera- tive protein, antibiotics to control microbial contamination, ture, so that stability is enhanced by cooling (30). Therefore, and one or more buffers to control the pH. Isotonicity, viral specimens generally should not be stored at room however, is not an absolute requirement; some highly suc- temperature or incubator temperature when a refrigerator is cessful transport media contain hypertonic solutions of available. The probability of a successful isolation is in- sucrose. Liquid transport media are used primarily for creased when the time interval between collection and transporting swabs or materials released into the medium inoculation of the culture is not prolonged and when the from a collection swab. Liquid media may be added to other specimen contains as many virus particles as possible. To specimens when inactivation of the viral agent is likely and maximize the amount of virus in the specimen, it should be when the resultant dilution is acceptable. collected early in the patient's illness. If a suitable transport The specimen should be collected as soon as possible after medium is used, successful and reliable isolations can be the onset of symptoms, preferably within 3 to 7 days (42). accomplished when viral specimens are sent to either a local The specimen contains the maximum amount of virus at the laboratory or a reference laboratory. time of collection. To maintain viral infectivity, the speci- A transport medium is not always essential for virus men should be placed in a good transport medium. 120 VOL. 3, 1990 TRANSPORT OF VIRAL SPECIMENS 121 Swabs Huntoon et al. and Smith (34, 61). The double-blind prospec- tive study was carried out to compare the rates of virus For transport of viral specimens, the swab may play one of recovery from the upper respiratory tracts of children, using two possible roles. First, it might be both the collection the three different transport media. The media were pack- device and the vehicle of transport to the laboratory or to the aged in the Culturette ampoules and labeled in code by the cell culture or to both. Second, the swab may function solely manufacturer. Each specimen was collected on the three as a collection device upon which the virus remains only as different swabs, and one each was inserted into the three long as it takes to deposit the swab in a tube of transport different Culturette systems, which were transported to the medium. In this case, another parameter that must be laboratory at room temperature in 30 min to 21 h for considered is whether the swab is to remain in or be removed inoculation into cell cultures. Specimens were collected from the transport medium after dispersal of the specimen. during all four seasons of the year. Eighty isolates were The materials in the swab or the swab shaft may be toxic to obtained from 200 specimens for a positive rate of 40%. the virus in the specimen. Viruses recovered were parainfluenza virus, enterovirus, The toxicity of calcium alginate for HSV was reported by adenovirus, HSV, influenza virus types A and B, respiratory Crane and associates in 1980 (18). Swab materials shredded syncytial virus (RSV), varicella-zoster virus (VZV), and from rayon, cotton, Dacron, polyester, and calcium alginate rhinovirus. Of the isolates, 90% were recovered in Hanks swabs were mixed with virus (HSV type 2 [HSV-2]) in BSS, 80% were recovered in Stuart, and 79% were recov- suspension. The suspensions were agitated with a magnetic ered in Leibovitz-Emory media, but the differences were not stirrer. At intervals up to 72 h, samples were removed for statistically significant. virus titration. Over this period of time the virus titers from A four-site evaluation of a laboratory test kit for rapid the samples exposed to rayon, cotton, Dacron, and polyester diagnosis of HSV infections was reported by Smith (62). All dropped a little more than 10-fold, an amount equal to the four laboratories processed specimens sent to their specific virus control, which was exposed to no swab material. In laboratories with the request for HSV culture. One labora- contrast, the virus exposed to calcium alginate dropped tory used the Culturette for transport, one used Hanks BSS more than 1,000-fold in 48 h. In follow-up experiments, these containing 0.5% gelatin, one used Eagle minimal essential authors showed that calcium alginate binds directly to the medium (MEM) with 2% bovine serum, and the fourth used HSV virion, rendering it noninfectious. Thus, for the collec- veal infusion broth with 0.5% gelatin. The transport times tion of herpesviruses, calcium alginate swabs should be were approximately 24 h or less. The recoveries of HSV avoided. from the specimens were essentially comparable. Direct Another study examining the effect of swab materials on comparison could not be made because the same specimen the recovery of HSV from clinical specimens (9) compared was not tested in all transport media. Yet, the Culturette had cotton swabs with calcium alginate swabs. There was no the highest recovery rate, suggesting that the inclusion of difference in the recovery of virus from cotton or calcium protein in the other three media was not a factor that alginate swabs when the samples