UK SMI B 4 issue 7.2 (August 2022): investigation of superficial mouth samples – GOV.UK

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Updated 20 September 2022

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Public Health England. ‘Investigation of Superficial Mouth Samples. UK Standards for Microbiology Investigations’ 2015: B 4 Issue 7
This SMI describes the processing and bacteriological and mycological investigation of superficial mouth samples. Predominately mouth swabs but saliva and oral rinses are also covered. Infections of salivary glands (parotid, submandibular and sublingual) include bacterial and viral infections and are not covered in this SMI.
This SMI should be used in conjunction with other SMIs.
Infections of the oral mucosa usually present as acute conditions. Usually, these arise from the colonising oral flora but can also result from a flare-up of a chronic low-grade infection.
Oral mucosal infections are typically associated with biofilms formed on the inanimate surfaces present in the oral cavity such as the teeth and dentures.
Infections of the gingiva (gingivitis, including acute ulcerative gingivitis) and periodontal tissues (periodontitis) are the most common forms of oral infection and processing specimens from these infections are covered in UK SMI B 14: investigation of pus and exudates.
Oral mucositis is a painful complication of chemotherapy or head and neck radiotherapy, caused by direct cytotoxicity of the treatment regime. Super-infection usually with yeasts and oral bacteria can exacerbate the problem and microbiological examination can help to guide symptomatic treatment.
Erythematous and pseudomembranous candidosis (1, 2) are the most frequent clinical presentations of oral fungal infection. The infections may involve the mucosal surfaces of the cheeks, tongue (dorsal and ventral surfaces) and both hard and soft palates. The most common cause is Candida albicans. Candida species other than C. albicans such as Candida glabrata may also be isolated, either alone or in combination with C. albicans. This is especially common in the medically compromised or those with a history of prolonged antifungal therapy (3, 4).
Atrophic candidosis (denture stomatitis) may occur in the palatal mucosa below the fitting surface of dentures, especially when patients sleep with their dentures in place and/or have xerostomia. Candida species other than C. albicans are important to identify since they may demonstrate reduced susceptibility and clinical resistance to the first-line anti-fungal agents and may be responsible for refractory or recurrent infections. Rarely, moulds may colonize and infect sinuses and result in palatal erosion. Specimens in the form of an oral rinse (known volume of sterile saline) are used to quantitatively determine colonisation or infection (5).
Angular cheilitis and peri-oral infections are common infections affecting the angles of the mouth and lips, usually caused by an intra-oral reservoir of infection, typically biofilms associated with denture stomatitis. Infection may be due to S. aureus, Candida species and/or group A streptococci. It is common for dentate patients with angular cheilitis to have infection with both S. aureus and C. albicans in the labial commissure region. Swabs should be taken from the lesions themselves. Swabs should also be collected from relevant intra-oral sites, for example, denture-fitting surface and the anterior nares to identify sites of colonisation to be treated with eradication therapy, to reduce relapse rates.
Patients who are severely medically compromised and have reduced salivary flow, together with parenteral feeding, may develop staphylococcal mucositis caused by S. aureus. Enterobacteria may also play a role in severe cases. The erythematous changes in the oral mucosa may be indistinguishable clinically from candidosis, requiring the need for microbiological investigation. Results should be interpreted in a clinical context since asymptomatic carriage of S. aureus or Enterobacteria may occur. Strict regular oral hygiene measures are usually sufficient to resolve clinical symptoms. Systemic antibiotics are not usually required although may play an important role in the management of severe oral mucositis in some patient groups such as the terminally ill (6).
There are many non-infective causes of oral ulceration such as traumatic ulcers, recurrent aphthous ulcers, inflammatory conditions and malignant lesions. Infective causes of oral ulceration are commonly viral in origin (for example, Herpes simplex). Uncommon bacterial causes of ulceration are syphilis and tuberculosis whilst other rare causes of oral ulceration include fungal infections such as histoplasmosis.
Abscess and deep-seated infections (dental abscesses, and salivary gland abscesses) are dealt with in B 14 – Investigation of abscesses and deep-seated wound infections.
Osteomyelitis, including bacterial, mycobacterial and fungal osteomyelitis are dealt with in B 42 – Investigation of bone and soft tissue associated with osteomyelitis.
Borrelia vincentii and Fusobacterium species are associated with the infection known as Vincent’s angina. It is characterised by ulceration of the pharynx or gums and occurs in adults with poor mouth hygiene or serious systemic disease (7), B 9 – Investigation of throat-related specimens provides more information.
The recommendations made in UK SMIs are based on evidence (for example, sensitivity and specificity) where available, expert opinion and pragmatism, with consideration, also being given to available resources. Laboratories should take account of local requirements and undertake additional investigations where appropriate. Prior to use, laboratories should ensure that all commercial and in-house tests have been validated and are fit for purpose.
Selective media which does not support the growth of all circulating strains of organisms may be recommended based on the evidence available. A balance therefore must be sought between available evidence, and available resources required if more than one media plate is used.
SMIs use the term ‘CE marked leakproof container’ to describe containers bearing the CE marking used for the collection and transport of clinical specimens (8, 9). The requirements for specimen containers are given in the EU in vitro Diagnostic Medical Devices Directive (98/79/EC Annex 1 B 2.1) which states: “The design must allow easy handling and, where necessary, reduce as far as possible contamination of and leakage from, the device during use and, in the case of specimen receptacles, the risk of contamination of the specimen. The manufacturing processes must be appropriate for these purposes”.
This section covers specific safety considerations related to this UK SMI (8 to 24)
Use aseptic technique.
Mouth swab, saliva and oral rinse
For safety considerations refer to section 5.1.
Collect specimens before starting antimicrobial therapy where possible (26).
To assure that the preconditions of the sampling for oral infections are comparable it is advised that patients should not:
If possible samples should be taken in the morning under fasting conditions.
Unless otherwise indicated collect each swab for bacterial and/or fungal culture and place in appropriate transport medium (25, 27, 28, 29, 30).
Collect specimens other than swabs into appropriate CE-marked leakproof containers and place in sealed plastic bags.
Sample any lesions or inflamed areas using cotton-tipped swabs. Samples of denture fitting surfaces should also be swabbed as these are more sensitive sites than the palatal mucosa to recover Candida species. The use of a tongue depressor or spatula may be helpful. Oral rinses can be useful to follow up level of colonisation. These are collected by rinsing with 10ml of sterile saline for one minute.
Numbers and frequency of specimens collected depend on the clinical condition of patient (26).
For safety considerations refer to section 5.1.
Specimen processing and procedure (8, 9).
Most mouth samples are swabs unless the patient is immunocompromised or has other clinical indications.
Saliva samples may be collected for microbiological investigation and for other types of assessment. Increasingly saliva is being used as a sample for new diagnostic techniques, but also for assessing xerostomia and risk of dental caries. Care is needed to avoid contamination of these specimens and cross infection from these specimens. Sometimes culture is done with an exact volume of saliva in order to assess the count of a particular organism (for example, S. mutans or lactobacilli per ml of the original saliva sample.
Not applicable.
For safety considerations refer to section 5.2.
For comparison it is sometimes useful to dilute neat sample 1:100 (0.1ml + 9.9ml PBS). Inoculate 50µl onto a Columbia Blood Agar and use a hockey stick to spread out. A MacConkey/Cystine lactose electrolyte deficient agar (CLED) plate may also be useful.
Direct microscopic examination with Calcofluor staining may be helpful if histoplasma or mould infection is suspected.
Inoculate each agar plate using a sterile loop or a loopful of liquid (Q 5 – Inoculation of culture media for bacteriology).
For the isolation of individual colonies, spread inoculum with a sterile loop.
If histoplasmosis is suspected the length of incubation should be extended and carried out in Containment Level 3.
If an unusual fungal infection is suspected a second Sabouraud plate should be set up at 30°C and incubation time extended.
If an unusual fungal infection is suspected a second Sabouraud plate should be set up at 30°C and incubation time extended.
Refer to individual SMIs for organism identification.
Organisms may be further identified if this is clinically or epidemiologically indicated.
Refer to British Society for Antimicrobial Chemotherapy (BSAC) and/or EUCAST guidelines.
C. albicans is not routinely tested unless associated with recurrent infection, requested by clinician or the patient’s history indicates significant immunosuppression.
Not applicable.
‘Specialist and reference microbiology: laboratory tests and services’ provides information on the tests offered, turnaround times, transport procedure and the other requirements of the reference laboratory.
Organisms with unusual or unexpected resistance, or associated with a laboratory or clinical problem, or anomaly that requires elucidation should be sent to the appropriate reference laboratory.
Contact the appropriate devolved national reference laboratory for information on the tests available, turnaround times, transport procedures, and any other requirements for sample submission.
Report for fungi if applicable.
All results should be issued to the requesting clinician as soon as they become available unless specific alternative arrangements have been made with the requestors.
Urgent results should be telephoned or transmitted electronically in accordance with local policies.
Report clinically significant organisms isolated or report:
For rinses, report as (5):
Interim or preliminary results should be issued on detection of potentially clinically significant isolates as soon as growth is detected, unless specific alternative arrangements have been made with the requestors.
Urgent results should be telephoned or transmitted electronically in accordance with local policies. 
Final written or computer-generated reports should follow preliminary and verbal reports as soon as possible.
Report susceptibilities as clinically indicated. Prudent use of antimicrobials according to local and national protocols is recommended.
The Health Protection (Notification) regulations 2010 require diagnostic laboratories to notify UKHSA (31 to 36) when they identify the causative agents that are listed in Schedule 2 of the Regulations. Notifications must be provided in writing, on paper or electronically, within 7 days. Urgent cases should be notified orally and as soon as possible, recommended within 24 hours. These should be followed up by written notification within 7 days.
For the purposes of the Notification Regulations, the recipient of laboratory notifications is the local UKHSA Health Protection Team. If a case has already been notified by a registered medical practitioner, the diagnostic laboratory is still required to notify the case if they identify any evidence of an infection caused by a notifiable causative agent.
Notification under the Health Protection (Notification) Regulations 2010 does not replace voluntary reporting to UKHSA. The vast majority of NHS laboratories voluntarily report a wide range of laboratory diagnoses of causative agents to UKHSA and many UKHSA Health protection Teams have agreements with local laboratories for urgent reporting of some infections. This should continue.
Note: The Health Protection Legislation Guidance (2010) includes reporting of Human Immunodeficiency Virus (HIV) and Sexually Transmitted Infections (STIs), Healthcare-Associated Infections (HCAIs) and Creutzfeldt–Jakob disease (CJD) under ‘Notification Duties of Registered Medical Practitioners’: it is not noted under ‘Notification Duties of Diagnostic Laboratories’.
Other arrangements exist in Scotland (33, 34), Wales (35) and Northern Ireland (36).
For a text equivalent of this flowchart, see below.
If histoplasmosis is suspected the length of incubation should be extended and carried out in Category 3 conditions.
If an unusual fungal infection is suspected a second Sabouraud plate should be set up at 30°C and incubation time extended.
The specimen can follow 1. the standard media pathway or 2. the supplementary media pathways.
Specimens should be cultured onto Sabouraud agar, incubated at 35 to 37°C in air for 40 hours or onto blood agar incubated at 35 to 37°C in 5 to 10% CO2 for 24 hours. (If Histoplasmosis is suspected on Sabouraud agar the length of incubation should be extended and carried out in Category 3 conditions.)
If an unusual fungal infection is suspected a second Sabouraud plate should be set up at 30°C and incubation time extended.
Results on Sabouraud agar indicate C. albicans, non albicans yeast.
Results on blood agar indicate Group A strep, S. aureus and coliforms.
Specimens should be cultured onto MacConkey or CLED agar and incubated at 35 to 37’C in air for 24 hours.
Results indicate coliforms, non fermentative Gram negative.
1. Rautemaa R and Ramage G. ‘Oral candidosis–clinical challenges of a biofilm disease.’ Critical Reviews in Microbiology 2011: volume 37, pages 328 to 336
2. Bagg J and others. ‘High prevalence of non-albicans yeasts and detection of anti-fungal resistance in the oral flora of patients with advanced cancer.’ Palliative Medicine 2003: volume 17, pages 477 to 481
3. Bagg J and Sweeney MP. ‘Oral problems in advanced cancer.’ CME Cancer Medicine 2003: volume 2, pages 23 to 28
4. Jobbins J and others. ‘Oral and dental disease in terminally ill cancer patients.’ British Medical Journal 1992: volume 304, page 1,612
5. Epstein JB and others. ‘Quantitative relationships between Candida albicans in saliva and the clinical status of human subjects.’ Journal of Clinical Microbiology 1980: volume 12, pages 475 to 476
6. Jackson MS and others. ‘Staphylococci in the oral flora of healthy children and those receiving treatment for malignant disease.’ Microbial Ecology in Health and Disease 2000: volume 12, pages 60 to 64
7. Finegold S. ‘Anaerobic Gram-Negative Rods: Bacteroides, Prevotella, Porphyomonas, Fusobacterium, Bilophila, Sutterella’. In: Gorbach SL, Bartlett JG, Blacklow NR, editors. Infectious Diseases, second edition. Philadelphia: WB Saunders Company 1998: pages 1,904 to 1,915
8. European Parliament. UK Standards for Microbiology Investigations (SMIs) use the term “CE marked leak proof container” to describe containers bearing the CE marking used for the collection and transport of clinical specimens. The requirements for specimen containers are given in the EU in vitro Diagnostic Medical Devices Directive (98/79/EC Annex 1 B 2.1) which states: “The design must allow easy handling and, where necessary, reduce as far as possible contamination of, and leakage from, the device during use and, in the case of specimen receptacles, the risk of contamination of the specimen. The manufacturing processes must be appropriate for these purposes”.
9. Official Journal of the European Communities. ‘Directive 98/79/EC of the European Parliament and of the Council of 27 October 1998 on in vitro diagnostic medical devices’. 7 December 1998, pages 1 to 37
10. Health and Safety Executive. ‘Safe use of pneumatic air tube transport systems for pathology specimens’. 9/99
11. Department for Transport. ‘Transport of Infectious Substances’ 2011 Revision 5. 2011
12. World Health Organization. ‘Guidance on regulations for the Transport of Infectious Substances 2013 to 2014.’ 2012
13. Home Office. Anti-terrorism, Crime and Security Act. 2001 (as amended)
14. Advisory Committee on Dangerous Pathogens. ‘The Approved List of Biological Agents. Health and Safety Executive.’ 2013, pages 1 to 32
15. Advisory Committee on Dangerous Pathogens. Infections at work: Controlling the risks. Her Majesty’s Stationery Office. 2003
16. Advisory Committee on Dangerous Pathogens. Biological agents: Managing the risks in laboratories and healthcare premises. Health and Safety Executive. 2005
17. Advisory Committee on Dangerous Pathogens. Biological Agents: Managing the Risks in Laboratories and Healthcare Premises. Appendix 1.2 Transport of Infectious Substances – Revision. Health and Safety Executive. 2008
18. Centers for Disease Control and Prevention. Guidelines for Safe Work Practices in Human and Animal Medical Diagnostic Laboratories. MMWR Surveillance Summary 2012: volume 61, pages 1 to 102
19. Health and Safety Executive. ‘Control of Substances Hazardous to Health Regulations. The Control of Substances Hazardous to Health Regulations 2002, fifth edition.’ HSE Books; 2002
20. Health and Safety Executive. ‘Five Steps to Risk Assessment: A Step by Step Guide to a Safer and Healthier Workplace.’ HSE Books. 2002
21. Health and Safety Executive. ‘A Guide to Risk Assessment Requirements: Common Provisions in Health and Safety Law.’ HSE Books. 2002
22. Health Services Advisory Committee. ‘Safe Working and the Prevention of Infection in Clinical Laboratories and Similar Facilities.’ HSE Books. 2003
23. British Standards Institution (BSI). ‘BS EN12469 – Biotechnology – performance criteria for microbiological safety cabinets.’ 2000
24. BSI. ‘BS 5726:2005 – Microbiological safety cabinets. Information to be supplied by the purchaser and to the vendor and to the installer, and siting and use of cabinets. Recommendations and guidance.’ 24 March 2005, pages 1 to 14
25. Barber S and others. ‘Evaluation of bacteriological transport swabs’ Pathology 1998: volume 30, pages 179 to182
26. Baron EJ and others. ‘A guide to utilization of the microbiology laboratory for diagnosis of infectious diseases: 2013 Recommendations by the Infectious Diseases Society of America (IDSA) and the American Society for Microbiology (ASM).’ Clinical Infectious Diseases 2013: volume 57, E22 to E121
27. Rishmawi N and others. ‘Survival of fastidious and nonfastidious aerobic bacteria in three bacterial transport swab systems.’ Journal of Clinical Microbiology 2007: volume 45, pages 1,278 to 1,283
28. Van Horn KG and others. ‘Comparison of the Copan ESwab system with two Amies agar swab transport systems for maintenance of microorganism viability.’ Journal of Clinical Microbiology 2008: volume 46, pages 1,655 to 1,658
29. Nys S and others. ‘Comparison of Copan eSwab with the Copan Venturi Transystem for the quantitative survival of Escherichia coli, Streptococcus agalactiae and Candida albicans.’ European Journal of Clinical Microbiology and Infectious Diseases 2010: volume 29, pages 453 to 456
30. Tano E and Melhus A. ‘Evaluation of three swab transport systems for the maintenance of clinically important bacteria in simulated mono- and polymicrobial samples.’ APMIS 2011: volume 119, pages 198 to 203
31. Public Health England. ‘Laboratory Reporting to Public Health England: A Guide for Diagnostic Laboratories.’ 2013, pages 1 to 37
32. Department of Health. ‘Health Protection Legislation (England) Guidance’ 2010, pages 1 to 112
33. Scottish Government. ‘Public Health (Scotland) Act. 2008 (as amended)’
34. Scottish Government. ‘Public Health etc. (Scotland) Act 2008. Implementation of Part 2: Notifiable Diseases, Organisms and Health Risk States’. 2009
35. The Welsh Assembly Government. ‘Health Protection Legislation (Wales) Guidance’. 2010
36. Home Office. ‘Public Health Act (Northern Ireland) 1967 Chapter 36. 1967 (as amended)’
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