Penanganan Spesimen Swab untuk Pemeriksaan

SARS CoV-2

Basti Andriyoko
Pokja Molekuler PDS PatKLIn
KSM / Laboratorium Patologi Klinik RSUP Dr. Hasan Sadikin Bandung
Pokok
Bahasan

Potensi kesalahan praanalitik

Interfering substances (Inhibitor PCR)

Pemilihan Jenis Swab & Media Transport

Waktu Pengambilan Spesimen

Penyimpanan Spesimen

Pengiriman Spesimen

Penanganan Limbah
 Preanalytical Error

Lack of identification/misidentification
Inadequate procedures for specimen
(swab) collection
Presence of interfering substances
Collection of inappropriate or inadequate
material for quality or volume
Testing carried out outside of the
diagnostic window
Inadequate procedures for specimen
handling, transport, storage
 Interfering Substances / PCR Inhibitors

Schematic presentation of the attack points of polymerase

chain reaction (PCR) inhibitors during sample preparation and PCR.
During sample processing and extraction:
1. Nucleic acids may interfere with surfaces of the vessels
2. Substances may react with nucleic acids

During PCR: Bessetti J. An introduction to PCR inhibitors. 2007

3. Inhibit reverse transcription
4. Degrade or modify the template DNA
• Daya inhibisi tergantung konsentrasi
5. Annealing of primers to the template can be hampered
6. DNA polymerase is degraded inhibited or altered inhibitor
7. Substances may interfere with binding of probes or with their • Umumnya kit ekstraksi sudah bisa
fluorophores menghilangkan zat zat inhibitor
• Cek Internal control setiap sampel untuk
kemungkinan inhibitor → hasil invalid
 Mengenal Jenis Swab
Fiber Fiber
wrapped flocked
swab swab

Homogenisasi
lebih mudah
Fiber Type
Fiber Type
- organic: cotton
- Nylon
- Synthetic:
Rayon, Dacron
(Polyester)

Fiber Fiber
wrapped flocked
Sumber:www.copanusa.com
swab swab
Memilih Swab untuk Pengambilan spesimen
Swab: Plastic aplicator Hindari penggunaan
Flocked swab lebih baik material organik

www.pathlab.co.nz

www.cell-nest.com

Kualitas Bahan Swab mempengaruhi hasil PCR

www.bd.com

Cek sertifikasi : FDA, CE, ISO

CEK SWAB SEBELUM DIGUNAKAN
Transport Medium

Viral / Universal Transport Medium

VTM / • Broth, BSA (bovine serum albumin), antibiotik
UTM • Menjaga viabilitas virus
• Untuk Kultur dan molekuler

Inactivated Transport Media

• Guanidine thiocyanate) or molecular grade Ethanol
ITM • Inactivate microbial viability (risk biohazard menurun)
• Preserved RNA and DNA in the sample
• Untuk pemeriksaan molekuler

NaCl • NaCl 0.9% / PBS

• Beberapa ada yang ditambahkan RNAse inhibitor
/ PBS • Untuk molekuler, kultur harus segera
 ITM (Inactivated Transport Media)

VIRAL TRANSPORTATION IN COVID-19 PANDEMIC: INACTIVATED VIRUS

TRANSPORTATION SHOULD BE IMPLEMENTED FOR SAFE TRANSPORTATION AND
HANDLING AT DIAGNOSTICS LABORATORIES
Rajan Dewar, et al

• A variety of transport media have previously been shown to effectively inactivate/kill viral,
bacterial and fungal pathogens while preserving stability of the released DNA and RNA for
diagnosis. These media include ones with a surfactant (e.g. Guanidine thiocyanate) or
molecular grade Ethanol.
• Moving to virus-inactivating VTM at collection allows risk mitigation from transportation and
handling of bio-specimens for diagnosis and can potentially reduce the need for special
packaging and transportation measures for SARS-CoV-2/COVID-19 test samples.
 CEK MEDIA TRANSPORT SEBELUM DIGUNAKAN
CEK CAIRAN MEDIA:
- WARNA
- KEJERNIHAN
- VOLUME

Pertumbuhan Volume
jamur cukup

Kosong
Bocor
 Waktu Pengambilan Spesimen

Nasopharyngeal and oropharyngeal swabs

were collected and placed in the same tube
as URT specimen, and sputum was used as
LRT specimen

Lippi et al.: Vulnerabilities in the diagnosis of COVID-19

Kemungkinan positif pada pengambilan
swab ke 3 saat diagnosis

A.T. Xiao, et al. Dynamic profile of RT-PCRfindings from 301 COVID-19 patients in
Wuhan,China: A descriptive study. Journal of Clinical Virology 127 (2020)
Penyimpanan Spesimen

72 jam

48 jam

Repeated freezing and thawing of specimens must be avoided

March 2012 Volume
50 Number 3

Temperature: 40C Room Temperature

Temperature: 370C
The types of swabs were as follows:
- plain (rayon-tipped) swabs without medium
- virus transport medium swabs in a medium-soaked
sponge (VTM)
- flocked swabs with universal transport medium (UTM)
- flocked nylon fiber swabs with liquid Amies medium
(E swabs)
- swabs with Amies gel containing charcoal
 Pengiriman Spesimen

• Triple layer packaging

• Suhu pengiriman +40C
 Keamanan dalam Pengiriman Sampel

Primary
tube Absorbent &
Secondary
container
Rigid outer packaging
Pengiriman Internal Faskes
 Penanganan Limbah Infeksius

Solid waste should be sterilized by:

- high pressure steam heat (in autoclave of
sufficient size that permits the adequate
steam flow and with physical or biological
indicators that ensure effectiveness of the
process)
- or directly incinerated (in conventional Autoclave :
double chamber incinerator). 1210C, 30 menit Incinerator
COVID-19 WASTE MANAGEMENT FACTSHEET
 Kesimpulan
Pengambilan saat awal onset
gejala

Flocked swab lebih baik

Untuk PCR ITM lebih aman

Pastikan suhu penyimpanan sesuai

Prinsip: Triple layer packaging

Autoclave / Incinerator
TERIMA
KASIH
Pustaka
• PCR inhibitors – occurrence, properties and removal
https://sfamjournals.onlinelibrary.wiley.com/doi/epdf/10.1111/j.1365-2672.2012.05384.x
• An Introduction to PCR Inhibitors
https://www.researchgate.net/publication/285475103_An_introduction_to_PCR_inhibitors
• Potential preanalytical and analytical vulnerabilities in the laboratory diagnosis of coronavirus disease 2019 (COVID-19)
https://www.degruyter.com/view/journals/cclm/58/7/article-p1070.xml?tab_body=pdf-78264
• How to choose your waste management technology to treat COVID-19 waste
https://wedocs.unep.org/bitstream/handle/20.500.11822/32778/FS3.pdf?sequence=1&isAllowed=y
• Collecting, preserving and shipping specimens for the diagnosis of avian influenza A(H5N1) virus infection. Guide for field
operations
https://www.who.int/ihr/publications/CDS_EPR_ARO_2006_1.pdf?ua=1
• Evaluation of Swabs, Transport Media, and Specimen Transport Conditions for Optimal Detection of Viruses by PCR
https://jcm.asm.org/content/jcm/50/3/1064.full.pdf
• VIRAL TRANSPORTATION IN COVID-19 PANDEMIC: INACTIVATED VIRUS TRANSPORTATION SHOULD BE IMPLEMENTED FOR
SAFE TRANSPORTATION AND HANDLING AT DIAGNOSTICS LABORATORIES
https://www.archivesofpathology.org/doi/pdf/10.5858/arpa.2020-0175-LE
• Dynamic profile of RT-PCR findings from 301 COVID-19 patients in Wuhan, China: A descriptive study
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7151472/pdf/main.pdf
• Coronavirus disease 2019 (COVID-19) Quick Reference for Clinical Health Care Workers
https://www.nicd.ac.za/wp-content/uploads/2020/05/COVID-19-Quick-reference-v13-15.05.2020.pdf
• GENERAL PROCEDURES FOR INACTIVATION OF POTENTIALLY INFECTIOUS SAMPLES WITH EBOLA VIRUS AND OTHER
HIGHLY PATHOGENIC VIRAL AGENTS
https://www.paho.org/hq/dmdocuments/2014/2014-cha-procedures-inactivation-ebola.pdf