Viral Specimens: Collection, Transportation and Preservation for Laboratory Diagnosis of Viruses

Viral Specimens- Collection, Transportation and Preservation for Laboratory Diagnosis of Viruses

Viral Specimens collection

Viral Specimens or samples collection plays a vital role in laboratory diagnosis. Proper sample collection leads to the proper diagnosis of disease. Sample should be collected aseptically. Appropriate samples from the appropriate site and an adequate amount should be collected. Once collected samples are recommended to emulsify or mixed properly with viral transport medium (VTM).

Collection and Transport of Virological Specimens 

In the investigation of viral infections, reliable laboratory results can only be obtained if the specimens are collected, preserved, and transported correctly to the Virology laboratory.

When to collect viral specimens?

  • Specimen for detection of viruses should be collected as soon as possible after the appearance of the symptoms that is when the concentration of the virus is at its highest.
  • Blood for antibody testing should be collected as early in the disease as possible and also 5- 10 days later.
  • Two serum specimens (paired sera) are necessary to detect a rise in the titer of the Ab. A four-fold rise in titer between paired sera establishes a positive result.

Central nervous system-

  • CSF

Respiratory system

Upper respiratory-

  • Throat swab
  • Nasopharyngeal swab
  • Ear discharge

Lower respiratory-

  • Sputum
  • Bronchoscopy specimen
  • Tracheostomy/endotracheal aspirate
  • Lung biopsy/aspirates

Eye –discharge

GIT

  • Stool
  • Rectal swab
  • Ascitic fluid

Genitourinary system

  • Urine
  • Vaginal/endocervical swab
  • Urethra discharge

Body fluids

  • Blood
  • Bile
  • Peritoneal effusion
  • Pleural effusion
  • Pericardial effusion
  • Synovial fluid

Surface specimen

  • Skin, nails, hair
  • Wound /ulcers swabs

Surgical specimen

  • Tissue biopsy
  • Abscesses

The following information is intended only as a general guide for viral sample collection-

Feces

  1. Place about 4-8 g of feces (small spoon) in a clean, dry, leak-proof container.
  2. Deliver to the laboratory as soon as possible.
  3. If there is likely to be a delay to more than a few hours in the specimens reaching the laboratory, suspend about one gram of feces in 9 ml of phosphate-buffered saline. If possible centrifuge at 2000 g for 15 minutes, and then transfer the supernatant fluid to a clean leak-proof container.
  4. Label and keep at -20 °C.
  5. Send to the virology laboratory in a cold box.
  6. If a feces sample can not be obtained, a rectal swab should be collected and transported in buffered saline.
  7. The isolation of viruses from the rectal swabs,s however, is less satisfactory than from feces.
  8. Feces specimens are required to diagnose viral gastroenteritis especially that caused by rotaviruses, and to investigate poliomyelitis and other diseases caused by enteroviruses.
Recommended specimens for viral diagnosis
Recommended specimens for viral diagnosis

Nasopharyngeal secretions

  1. Collect a specimen by passing a sterile cotton wool swab, attached to a pent piece of thin wire, through the floor of the nasal cavity as far as the nasopharynx.
  2. After few seconds, slowly withdraw the swab and immerse the swab in a container of sterile VTM, cutting off swab wire to allow the bottle cap to be replaced tightly.
  3. A specimen of nasopharyngeal secretion can also be obtained by using a sterile mucus extractor.
  4. Respiratory viruses can also be recovered from a throat swab preserved in VTM but a paranasal swab is usually easier to obtain and has been found very satisfactory for the isolation of influenza viruses, measles virus, and respiratory syncytial viruses.
  5. Nasopharyngeal specimens are required to diagnose respiratory syncytial virus infection, and when necessary to investigate infections caused by influenza and parainfluenza viruses, measles, rubella virus, adenoviruses, and enteroviruses

Cerebrospinal fluid

  • Collect 0.5- 1.0 ml of fluid in a dry, sterile, leak-proof container. Refrigerated immediately at 4°C.
  • Transport in an insolated cold box.
  • Cerebrospinal fluid is required to investigate some arbovirus infections. When meninges are infected, the CSF will contain lymphocytes and the CSF total protein will be raised.

Skin and ulcer specimens

  1. Collect skin scrapings in a dry sterile container.
  2. Refrigerate immediately at 4°C, and then transport in a cold box.
  3. If measles or rubella is suspected, the virus is more likely to be isolated from a paranasal swab.
  4. Skin specimens are required to investigate infections caused by herpes simplex viruses, rabies virus.

Blood for serological tests

  1. Two serum specimens are required to diagnose an infection serologically.
  2. Collect the sample within 5 days of the onset of the symptoms, and the second sample 5-10 days later.
  3. Collect 5-10 ml of venous blood in a dry sterile, screw-cup glass tube or bottle.
  4. A single sample is required for investigating newborn infants with congenital defects, determining immunity to rubella or other viruses.
  5. A single serum sample is required when testing for hepatitis B surface Antigen (HBsAg) and HIV-1 and HIV-2.
  6. After the blood is collected, collect it in a leak-proof container, refrigerate it at 4°C until transport in an ice box to the virology laboratory.

 Anticoagulated blood for culture

Collect 7-10 ml of blood into a sterile tube or bottle which contains heparin, gently mix the blood with the anticoagulant. Viruses which can be cultured from blood include dengue viruses and some arboviruses. Specimens such as urine and sputum are generally unsuitable for viruses’ isolation.

Information to accompany specimens

A request form should include the following information:

  1. Name, place, age, number of the patient, and details of any recent travel.
  2. Type of specimen, if two, mention as the first of second.
  3. Investigation required.
  4. Full clinical information: nature duration, and severity of the disease.
  5. Details about immunization and antimicrobial therapy.
  6. Name and address of the hospital or health centre.

Transporting of specimens to the Virology laboratory

  • Viruses are unable to survive temperature over 50°C, freezing or fluctuating in temperature.
  • Viruses can also be damaged be light, drying, change in pH, and bacterial enzymes.
  • The usage of VTM will prevent specimens from drying out and help to preserve viral activity.
  • All viral specimens should be transported in an icebox with a warning label.

Storage and Transportation of Viral specimens

Sooner the specimens reach the laboratory the better.

By lowering the temperature

  • Short term storage : +4°C
  • Long term storage: -70°C/-190°C(liquid nitrogen)

Use of Viral Transport medium

  • Specimens are emulsified in the medium

VTM (Viral Transport Medium)

  • Composed of:
  1. Buffered salt solution
  2. Proteins (Bovine serum albumin)
  3. Antimicrobial agents
  4. Indicator
  • Function
  1. Preserves viral infectivity within the specimen
  2. Prevents specimen from drying
  3. Prevents growth of bacteria and fungi

Criteria for rejecting Vial specimens

  • Mismatch of information on the label and the request
  • Inappropriate transport temperature
  • Excessive delay in transportation
  • Inappropriate transport medium
  1. specimen received in a fixative
  2. dry specimen
  • Insufficient quantity
  • Leakage

Key Notes on Viral Specimens

  • All samples which may contain dangerous and highly infectious viruses must be labeled HIGH RISK with a red marker pen, and handled with great care.
  • Term              Understanding definition
  1. Common cold: Respiratory infection by Rhino virus, Coronavirus, Para-influenza virus, Influenza virus
  2. Flu: Common cold caused by Influenza virus (only)
  3. Influenza: A virus that causes common flu or  specific flu (bird flu, swine flu)
  4. Swine flu: A type of Influenza virus transmitted to humans often causing severe respiratory disease
  5. Bird flu/ Avian Influenza: A type of influenza virus that infects birds and
  6. H5N1/ H1N1: Scientific name of a virus causing bird flu
  7. Pneumonia: Severe Infection by virus or bacteria in the lower part of our lungs
  8. SARS: Severe Acute Respiratory Syndrome, or severe respiratory disease by one type virus (coronavirus).
  9. SARS-CoV-2: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an appropriate name for the new coronavirus.

References

  1. https://path.upmc.edu/Virology/G-02.htm
  2. https://cmr.asm.org/content/cmr/3/2/120.full.pdf
  3. https://www.researchgate.net/publication/284875818_Specimen_collection_transport_and_processing_Virology
  4. https://www.laboratoryalliance.com/healthcare-providers/laboratory-services/specimen-collection-documents/virology-specimen-collection-guidelines/
  5. https://www.who.int/ihr/publications/CDS_EPR_ARO_2006_1.pdf
  6. https://www.who.int/influenza/rsv/rsv_collection_transport_storage_samples/en/
  7. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC358147/https://reallaboratory.com/product/virus-transport-and-preservation-medium-inactivated/
  8. https://www.cdc.gov/coronavirus/2019-ncov/lab/guidelines-clinical-specimens.html
  9. https://watermark.silverchair.com/22-5-766.pdf
  10. https://www.thelancet.com/journals/lancet/article/PIIS0140-6736(20)30557-2/fulltext
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