Collecting and transporting sputum specimens Module 3: Collecting and transporting sputum specimens Global Laboratory Initiative – Xpert MTB/RIF Training Package

World Health Organization Contents of this module 1 April, 2017 Choosing suitable specimen containers Timing of specimen collection How to collect a specimen Handling specimens and referring them for testing Labelling specimens

Learning objectives At the end of this module, you will be able to: Describe specifications of suitable containers for sputum collection Discuss different sputum collection strategies: spot/morning/spot, spot/spot, single spot for Xpert MTB/RIF Describe and demonstrate safe and correct collection of sputum Describe options for specimen collection, handling and transport List features of a good quality sputum specimen Outline requirements for a properly labelled specimen

Sputum-specimen collection: container specifications Adapt according to the NTP’s guidelines in your country 30-50 ml capacity Translucent or clear material Sides and walls that allow easy labelling Single-use combustible material Leak-proof with a screw-cap Wide mouth

Sputum specimen collection: timing World Health Organization 1 April, 2017 Sputum specimen collection: timing Spot–morning–spot WHO and The Union recommend this timing: Spot In the clinic – during the initial visit Early morning At home – first sputum produced in the morning on the day of the second visit to the clinic (ideally, the day after the initial visit) Spot In the clinic – additional sputum collected during the second visit.

Collection strategies: spot-morning-spot Advantages A spot specimen is available in case the patient does not return with the morning specimen. Disadvantages Requires two visits to the clinic. Diagnosis takes at least 2- 3 days. Increases the workload. There is a high risk of missing a case if only the first specimen is received.

Collection strategies: WHO’s revised policy WHO’s policy on case detection by microscopy was revised in 2007: The recommended number of specimens examined was reduced from three to two in countries with appropriate external quality assurance (EQA) procedures and documented good quality microscopy In these settings, the case definition was also revised to one AFB positive smear, defined as one or more acid-fast bacilli in at least 100 microscopic fields If appropriate quality assurance is not available, three specimens should be examined.

Same-day strategy: spot-spot Good quality microscopy of two consecutive sputum specimens (spot-spot) identifies 95–98% of smear- positive TB patients. Advantages – this strategy: Greatly reduces the workload of laboratories Has the potential for offering same-day diagnosis Is better for patients because it reduces the number of visits while largely maintaining sensitivity. Disadvantages: Very minor loss in the number of cases detected.

Sputum collection: overview Follow the NTP’s guidelines on sputum collection. Ensure that sputum collection is supervised. A single sputum specimen is recommended for Xpert MTB/RIF: Increased yield is obtained by testing multiple specimens but this increases the cost of testing An additional sputum specimen may be needed in case of an error or invalid Xpert MTB/RIF result At least 1 ml of sputum should be collected. Additional sputum specimens may be needed for microscopy, culture and DST, depending on the NTP’s guidelines (Xpert MTB/RIF is not recommended for monitoring patients’ treatment). Avoid using pooled sputum specimens.

The NTP’s guidelines for sputum collection in (country name) [Insert the NTP's specific guidelines] To be customized by each country

Sputum collection: safety When providing a sputum specimen, a patient may produce infectious aerosols and therefore biosafety precautions are needed: Instruct the patient to cover his or her mouth when coughing Never collect sputum in the laboratory Collect sputum away from other people in a well ventilated space following the NTP’s guidelines Do not stand in front of the patient during specimen collection!

Sputum collection: Patient education and instructions The best specimen comes from the lungs. Saliva or nasal secretions are unsatisfactory. Specimens should not contain food or other particles because the test may not work properly. Patients should be instructed to take the following steps to produce the best specimen: Wash your mouth with clean water to remove food and other particles Inhale deeply 2–3 times and breathe out strongly each time Cough deeply from your chest to produce sputum Place the open container close to your mouth to collect the specimen; do not get sputum on the outside of the container Wash your hands after collecting the sample.

Specimen quality: Optimal quality Mucoid Purulent (Images courtesy of A. Van Deun)

Specimen quality: Suboptimal quality Bloodstained Salivary (Images courtesy of A. Van Deun)

Specimen quality: overview Obtaining an adequate quantity of good quality sputum is critical to ensuring accurate test results. For best results, obtain 1-4 ml of purulent/mucoid sputum.

Request for sputum examination A request for sputum examination should include the: Name of the treatment unit Date of the request Patient's information (that is, name, sex, age, address and the register number of the patient or suspected case) Number of specimens and types of specimens sent for testing Date the specimens were collected Reason for examination (for example, diagnosis or follow-up) Signature of the person requesting the examination.

Laboratory request form (example) (1 of 2) To be customized by each country. Add your local request form, and show participants how to correctly complete the form.

Laboratory results form (example) (2 of 2) To be customized by each country. Add your local request form, and show participants how to correctly complete the form.

Labelling the specimen container Label with the patient’s name, identification number and the date of collection. Label the outer sides of the container with permanent ink. Never label the lid.

Receiving specimens in the laboratory Check the quantity and quality of specimens: Check the volume (ideally 1-4 ml; a minimum of 1 ml is required for Xpert MTB/RIF testing) Record sputum consistency (mucoid, purulent, bloody or watery) on the report form Ensure that specimens do not contain food or other particulates. Check that the patient’s information is complete, and that Xpert MTB/RIF testing has been requested in accordance with the NTP’s guidelines. Follow the NTP’s guidelines for rejecting samples. Ensure that the patient’s information is complete on both the request form and the specimen container, and ensure that the information matches. Record the specimen in the laboratory register and allocate a laboratory serial number to the specimen.

If patients are referred If specimens are referred Referral systems Peripheral facilities that do not perform Xpert MTB/RIF testing should refer either patients or specimens to a testing laboratory. If patients are referred If specimens are referred Advantages Sputum collection can be monitored and collection can be repeated if the first specimen is not satisfactory. Treatment can be initiated faster. Patient does not have to travel. Disadvantages Requires patients to travel. Requires systems for safe transport and shipment. Requires efficient system for transmitting results. Specimen transport may require cold-chain or preservatives.

Specimen referral The shipment of specimens constitutes a public-health risk, and the shipper is responsible for ensuring that samples are packed safely. Triple packaging (described in later slides) must be used: Locally appropriate solutions can be used as long as safety requirements are met. Country-specific safety guidelines must be followed in order to correctly categorize the specimen and its associated risk (eg, Category A or B, as per WHO Guidance on regulations for the transport of infectious substances, 2013–2014). To be customized by each country

Transporting specimens To be customized by each country Specimens should be transported to a laboratory as quickly as possible: A sputum specimen that has been delayed during transport can still be used for microscopy and Xpert MTB/RIF testing. Whenever possible, specimens should be refrigerated at 2–8°C for a maximum of 10 days. However, if necessary, specimens can be stored at a maximum of 35°C for up to 3 days, and then refrigerated at 2–8°C for a combined maximum duration of 10 days. The total number of specimens in a box must correspond to the number of accompanying request forms. The identification number on each sputum container must correspond to the number on a request form. The accompanying forms must contain the required information for each patient.

Triple packaging: Step 1 of 3 Primary packaging Wrap the leak-proof container in cotton wool or paper towels in a sufficient quantity to absorb the entire contents in case of leaks.

Triple packaging: Step 2 of 3 Secondary packaging Place the wrapped container inside a secondary container, such as a self- sealing plastic bag or another container. Place the secondary container in a rack to prevent leakage.

Triple packaging: Step 3 of 3 Tertiary packaging Place the secondary container and its contents in an approved safety cooler box or another appropriate container in an upright position. Place a biohazard sign – with markings and labelling appropriate for the specimen category – on the tertiary container.

Role play: Sputum collection Purposes To practise educating a person suspected of having TB about the importance of a properly collected sputum specimen and to practise providing instructions on how to collect a specimen To practice packaging sputum specimens for referral. Total time 40 minutes. Process Work in groups of 4. Choose roles: health worker, person suspected of having TB, dispatcher and observer. The health worker instructs the person suspected of having TB on how to produce a specimen, and then provides the specimen to the dispatcher for packaging; the observer provides feedback. Switch roles and repeat the process until everyone has played each part once.

Role play: Debrief Did the health-care worker follow appropriate procedures? COMPLETE a Request for Sputum Examination form LABEL the sputum containers EXPLAIN to the person suspected of having TB how to collect sputum and the importance of a properly collected specimen ASK the person suspected of having TB to produce a first sample ASK the person suspected of having TB to collect another sample the next morning and then bring it to the diagnostic centre (OR modify the advice to follow the NTP’s guidelines) PACKAGE appropriately the sputum sample for referral. Did the person suspected of having TB understand the instructions? Did the dispatcher receive the package appropriately and complete the administrative procedures correctly? What suggestions for improvements did the observer note?

Summary Not all containers are suitable for specimen collection: follow the NTP’s guidelines on the size or volume of the container to be used, and the material and type of lid. An adequate quantity (1-4 ml) of good quality sputum (that is, purulent/mucoid) as well as a properly completed Laboratory Request Form and properly labelled specimen containers are critical to ensuring accurate results from the Xpert MTB/RIF test. Patients should be educated on how to produce good quality specimens, and sputum collection should be supervised. A well functioning referral system (for patients or specimens, or both) is critical to ensuring a high quality diagnostic service: follow country-specific safety guidelines for storing and shipping samples; triple packaging must be always used.

Assessment What are the the NTP’s guidelines for collecting sputum specimens in your country? Why should sputum never be collected in a laboratory? What are the options for referral at peripheral health centres that do not perform the Xpert MTB/RIF test? What are the features of a good quality specimen? How should a sputum container be labelled? What supplies are needed for triple packaging for TB diagnostic samples?