MICROCOSM

Tumblr (Nov 2014) 218 followers

https://10minus6cosm.tumblr.com/

We started this blog in November 2014. The purpose is stated in the very first post.

“This blog was created to showcase the variety of micro-organisms encountered in the Diagnostic Bacteriology Laboratory at the Singapore General Hospital. It serves as a kind of digital museum of images (and anecdotes) which we hope will be of use to microbiology trainees, laboratory technologists, and appeal to a wider audience interested in microbiology.”

We had already realised over the years that SGH Microbiology Labs provide a rich resource of interesting micro-organisms and case scenarios (stories) that also have teaching value. To record them in one location saves us having to repeat these to different batches of residents or technologists and serves as a form of ‘institutional memory’. We purposely tried to write in a manner that laypeople would also be able to understand as we felt we should also create awareness and educate the general public about diagnostic clinical microbiology.

Social media platforms like this blog have put the power of publicising the lab achievements and capabilities in our own hands rather than having to rely on mainstream media or the official institutional PR apparatus. We were not too bothered about whether anyone would read what we put out, our first target audience was our own people and we felt that over time, the buildup of content would have its own value. So prospective residents can see from our cumulative posts that they will experience a rich training environment, (and any potential philanthropist, can get an inside view of what we do beyond a superficial lab visit). By controlling the narrative, we have been able to include behind-the-scenes thinking and activities that never come out in newspaper reports or journal articles. For example, see “Serious Group B Streptococcal Infection in Adults Associated with Eating Raw Fish: A New Paradigm”. If you only read newspaper or journal articles you would never have known how the novel association between raw fish and GBS infection was worked out in the first place.

Those of you who have been following us may be wondering why the posts are becoming more infrequent. First, most of the historical material we intended to put up has already been posted. Second, we became too busy to write new material, and third we started posting content that was primarily image-based onto another platform (of which more later).

Over the years, it has been gratifying to receive a little feedback that readers have found the posts informative and sometimes even useful. Now that we have a bit more time on our hands, we will look into adding content again on a regular basis. We apologise for not responding to queries in a timely fashion as we don’t monitor this blog assiduously.

Youtube (Nov 2014) 1.3K subscribers

https://m.youtube.com/@diagnosticbacteriology1796

We started a Youtube channel at the same time (November 2014) as that was the main platform for videos then. If you have not seen the tumbling motility of Listeria monocytogenes, or only read about aerial hyphae of Nocardia in text, go over and have a look.

Twitter (Mar 2015) 1.1K followers

https://x.com/10minus6cosm

We went on Twitter in March 2015 really just to find out what the buzz (then) was about. We subsequently found it was one of the best ways to get updated about developments in our fields of interest, and to get a sense of what was expert opinion around the world (especially in bioinformatics and information technology). This proved particularly useful during the chaos of the COVID-19 years. We don’t consider our own opinion as expert, so we mainly repost from people we follow, and highlight local microbiology news. Unfortunately most of the US experts have migrated from Twitter to Bluesky, though there is still enough content here to keep us on this platform. We are on Bluesky too, but mainly to stalk the American experts!

Facebook (March 2015) 1.3K followers

https://www.facebook.com/DeptMicrobiology/

We went on Facebook intending to use it as a notice board announcing department activities, and highlighting interesting articles (some our own). We also started cross-referring posts across our various social media platforms.

Instagram (Dec 2018) 606 followers

https://www.instagram.com/10minus6cosm/

This is our most active platform at present. It is easy to post material on Instagram and the word count is not restrictive. As a result it has become our main vehicle for image-based content with negative impact on our frequency of Tumblr posts. We make it a point to try and showcase more unusual microorganisms.

LinkedIn (Mar 2025) 37 followers

https://www.linkedin.com/company/sgh-department-of-microbiology/

We resisted going on LinkedIn for ages despite institutional encouragement because we saw it mainly as a jobs market with less broad international appeal than Facebook. Eventually we succumbed because more of our own hospital senior leaders are on LinkedIN!

WeChat (Sep 2025)

Well, this was a little experiment. We realised that none of the above social media platforms were visible in China, which was potentially a large audience. The primary target was our diagnostic microbiology counterparts there. We started on Weibo, but that seemed pretty dead, and quickly switched to WeChat. Basically we chose from our various social media posts (mainly Instagram), and reposted what we felt appropriate onto WeChat. This was only feasible because of the ability of A.I. (ChatGPT) to efficiently fine-tune Chinese translations from the original English text (we still got the translation vetted by a native mainland Chinese speaker).

We quickly discovered a few things. It is much more complicated getting onto a Chinese social media platform than a Western one (ChatGPT to the rescue again). Personal WeChat accounts come with a number of limitations including restrictions on posts that are visible to the public. You can get round this by starting an organisational account but that probably needs to be initiated by the hospital. Finally, we got the sense that on the whole it is not the culture of Chinese scientists/doctors to use social media for professional purposes. So far we have had no interest in our account.

Why are we on so many different platforms? The simple reason is each platform fulfils slightly different purposes. We also find the audiences often do not overlap. At present we find Instagram, Twitter, LinkedIn, and Facebook to be the best for outreach.

I am an MD microbiologist recently moved to Singapore and preparing for FRCPath part 2 exam. It will be a great help if you can suggest a study group or study partner for the same.

Part II preparation

I approached this quite differently from Part I - I didn't really use textbooks but instead spent a lot of time going through guidelines and websites instead.

I continued to use my Anki study deck from Part I (which functioned as revision for Part I knowledge), and kept adding to it during this time.

My study group prepared for this in a similar way to Part I viva by running through questions with each other – by the time we got to the exam we were all very used to answering questions over Zoom in the viva format!

Book reviews

How to be a lab director (I used a version that was around 2014, updated version is https://www.amazon.sg/How-Be-Lab-Director-2023/dp/B0BR9CYH1C/)

This is the one book that really stood out to me for RCPA Part II preparation

Very frank account of managing a laboratory that includes talking about management, quality and EQAs. But because it is so frank - some of it is stuff that you can't use as an answer during exams! Very useful for thinking about various aspects of lab management that you might not have come across previously.

Websites and guidelines (Australian)

Part I websites/guidelines/alerts remain important, though I went into the websites in far more detail this round and read a lot more guidelines.

Additional websites and guidelines that may be more Part II related than Part I related as below. I generally just went to each website and kept clicking on links and documents that sounded potentially related to microbiology then read as much as I could around these.

RCPA website

As for Part I - webinars, policies, guidelines, position statements

Also has slides/information on

Management and clinical governance

2022 workshop slides

Laboratory informatics

Pathology informatics workshops 2019 and 2023 slides https://www.rcpa.edu.au/Library/Practising-Pathology/PTIS/APUTS-FAQ

NPAAC/NATA

2020 risk based NPAAC standards

2022 NATA RCPA accreditation assessors workshop

POCT elements of a quality framework (June 2014)

ACSQHC https://www.safetyandquality.gov.au

All of the NPAAC documents Also has a lot of other documents that are related to microbiology, including

Antimicrobial stewardship

C diff surveillance

CARAlert

CLABSI surveillance

CPE control

Cumulative antibiogram

Environmental cleaning

Infection prevention

Mycobacterium chimaera

National antimicrobial resistance strategy

Open disclosure

Preventing and controlling infections standard

Sepsis

Bloodstream infection surveillance

Surgical site infection surveillance

Multi-resistant organism surveillance

Best to browse around the site and have a look at anything that sounds related to microbiology. The site is quite broad so need to sieve out microbiology-related things (like the above list) from the ones that are very unlikely to be of relevance to microbiology.

NATA/RCPA accreditation

NATA https://nata.com.au

Human Pathology ISO15189 https://nata.com.au/accreditation/medical-laboratory-accreditation-iso-15189/

There is also this gap analysis document https://nata.com.au/files/2023/03/SAG_11_2_ISO_15189_2022_Gap_Analysis.pdf that analyses the differences between 2012 and 2022 versions of ISO15189.

NATA accreditation process

https://www.odwyeraccreditation.com.au/nata-accreditation/nata-accreditation-process/

TGA https://www.tga.gov.au

This has a lot of information on medical devices (IVDs), biologicals, etc. Again probably best to browse around the site and have a look for microbiology-related things.

Some of the information they have that is likely relevant to microbiology include

IVD standards https://www.tga.gov.au/how-we-regulate/manufacturing/manufacture-medical-device/manufacture-specific-types-medical-devices/vitro-diagnostic-ivd-medical-devices

Classification of IVD medical devices

Biologicals regulation https://www.tga.gov.au/resources/resource/guidance/australian-regulatory-guidelines-biologicals-argb

Faecal microbiota transplant standards

Clinical performance requirements and risk mitigation strategies for HIV tests

Chlamydia/gonorrhoea/syphilis IVD self tests

COVID-19 in vitro diagnostics

COVID-19 rapid antigen tests

Hepatitis B/C viruses IVD self tests

Seasonal influenza rapid antigen combination tests

Clinical performance requirements and risk mitigation strategies for HIV tests

Regulatory requirements for in house IVDs

Fairwork Australia

Information about employment in Australia and issues surrounding this https://www.fairwork.gov.au/tools-and-resources/best-practice-guides/managing-underperformance

Australian Privacy Principles

https://www.oaic.gov.au/privacy/australian-privacy-principles/australian-privacy-principles-quick-reference

Individual state department of health websites, health.gov.au website

I went through the websites and clicked on anything that sounded potentially relevant to ID or microbiology. There is quite a bit of good information on these websites so is useful to just click through them, and some states have nice guidelines for certain things.

As for Part I – also looked at current health alerts here.

Clinical incident management

WA guideline 2019 https://www.health.wa.gov.au/~/media/Files/Corporate/Policy-Frameworks/Clinical-Governance-Safety-and-Quality/Policy/Clinical-Incident-Management-Policy-2019/Supporting/Clinical-Incident-Management-Guideline-2019.pdf

Queensland guide https://clinicalexcellence.qld.gov.au/sites/default/files/docs/resourses/clinicalincidentguide.pdf

Other useful sites and documents

NT guidelines for management of people with infectious diseases who put others at risk of infection

https://digitallibrary.health.nt.gov.au/prodjspui/bitstream/10137/976/1/Management%20of%20people%20with%20infectious%20diseases%20who%20knowingly%20put%20others%20at%20risk%20Guideline.pdf

Specific additional things to look up for Part II

Not comprehensive!

Business case proposal

New and emerging technologies

Total lab automation

Rapid blood culture diagnostics

AI in pathology

NGS and sequencing

New and emerging drugs e.g. olorofim, ibrexafungerp

OneHealth

Handling of risk group 4 agents

Workplace and policies - burnout, diversity/inclusion/equality, harassment/bullying, complaints policy

Privacy/confidentiality

Laboratory informatics and communication, including critical/high risk results

HIV testing algorithms

Outbreak investigation and management

Business continuity plan

Preparedness plan

Investigation of erroneous result/lab error

Introduction of new test

Quality system essentials

QC, QA, EQA, anything to do with quality

Centralisation/decentralisation of laboratories

Clinical governance pillars

Spill and hazard management

Laboratory safety

Risk management

Diagnostic stewardship

Deciding whether to send out a test or create an in house test

Molecular vs culture

Construction healthcare infection prevention

ISO15189

Open disclosure

POCT, self-testing, issues surrounding these

Additional notes about Part II preparation

I found it really useful to have definitions for various things e.g. privacy, confidentiality, bullying, harassment, discrimination, etc.

I also found it useful to have frameworks/templates for various topics

Outbreak investigation/management

Investigation of erroneous results, unexpected results, lab errors

Introduction of new test

Quality system essentials

Centralisation of laboratories

Dealing with staff problems

Clinical governance pillars

Spill/hazard management and risk

Business plan

Business continuity plan

Some buzzwords…

Multidisciplinary approach

Risk based approach

Risk vs benefit

Quality vs cost

Open disclosure

Our trainee Shireen recently passed her Royal College of Pathologists of Australasia Examination. She was invited to share her exam preparation strategy for the benefit of our other trainees.

Disclaimer

This document is based on my personal experience and what worked well for me to get me through the exams. Everyone’s experience will be different and what worked for me may not work so well for someone else (and vice versa)!

This is also not comprehensive and is just my attempt to cobble together a list of the resources that I used or found particularly helpful. There are also many other resources beyond the ones listed here!

Where possible I have tried to include links to various websites but please use the links at your own discretion.

It is also important to know your lab protocols well and know how the lab works.

Lastly don’t forget to go through the RCPA microbiology trainee handbook and the administrative requirements handbook!

General notes on exam preparation

Study tools

Anki flash cards – I am happy to share my Anki deck (with disclaimers)

I created my own Anki deck and slowly added to it over the course of about 3 years – had approximately 2400 cards by the time I took my Part II exam.

Every time I came across a new fact or something that I felt I needed to remember I would put the information into a flash card.

Anki has the advantage of being easily searchable, so the flash cards I made doubled as my study notes.

Anki syncs across all of my devices, so I created flash cards on my laptop and went through flash cards on my phone whenever I had a pocket of time (walking to/from the MRT, walking in the park, waiting for people).

Overall I found this an excellent tool to get my brain to remember microbiology trivia.

Study group

Having a study group made a massive difference to exam prep for me – I cannot overemphasise how helpful having one was!

Probably a good time to find a study group would be around a year before the Part I exam.

Question banks (not RCPA exam specific)

ID/micro MCQ book https://www.amazon.co.uk/Infectious-Diseases-Microbiology-Virology-Specialist/dp/1316609715/ - I went through this book before FRCPath I.

Jawetz has nice end of chapter MCQs.

Past year questions and question banks (RCPA exam specific)

RCPA question banks – RCPA publishes Part I Paper A questions for the last few years on the RCPA website

Ask your seniors if they have more questions – for example my study group made up practice questions for each other and therefore we have an additional self-generated question bank.

If I were to go back in time to about 1-1.5 years before my Part I, my approach would be to look through past year questions early to get a good idea of what sort of questions are being asked. Do as many practice questions as possible. Then see if can answer the questions for other similar organisms/illnesses too…

Start of training

I went through these books cover to cover. This is the approximate order in which I’d read these books, and I’d probably aim to finish these books in the first year or so of training

Book reviews

Clinical microbiology made ridiculously simple https://www.amazon.sg/Clinical-Microbiology-Made-Ridiculously-Simple-dp-1935660500/dp/1935660500/ This is a book targeted as medical students (I think) but I found this a great read at the start of training to get an overview and learn some mnemonics and ways to remember things that have stuck with me. Can get through this book fairly quickly!

Comprehensive review of infectious diseases https://www.amazon.com/Comprehensive-Review-Infectious-Diseases-Andrej/dp/0323568661/ Great introduction to ID and microbiology, but use with caution and some things may be different for our local setting/Australia

Murray (I used the 8th edition, updated edition is https://www.amazon.sg/Medical-Microbiology-Patrick-R-Murray/dp/) and/or Jawetz https://www.amazon.sg/Jawetz-Melnick-Adelbergs-Medical-Microbiology/dp/1260012026/ Both are undergraduate microbiology textbooks that cover fairly similar content at a fairly similar level. I read through both and preferred Murray for reading/learning, but liked the end of chapter MCQs in Jawetz.

Color Atlas of Medical Bacteriology (review is for third edition, updated edition is https://www.amazon.sg/Color-Atlas-Medical-Bacteriology-Luis/dp/1683670353/) Lots of nice pictures that show a lot of the bacterial isolates and biochemical reactions you need to become familiar with in the lab. Text also quite helpful as an introduction to each group of bacteria Fast facts bacteria chapter at the back is also very nice for quick reference/learning.

Part I preparation

Book reviews

Oxford handbook of Infectious Diseases and Microbiology https://www.amazon.sg/Oxford-Handbook-Infectious-Diseases-Microbiology/dp/019967132X/ This is unfortunately getting a bit dated now (2016), and is UK-based so some guidelines/methods are different from what we do here/in Australia. Can be a little dry (as it contains a lot of facts), but it has very nice chapters covering antimicrobials (sadly a bit outdated now) and infection control that can be read as introductions to these topics. Part 3 systematic microbiology contains many of the important organisms you need to know about by the time you take RCPA Part I. (Note this is not a comprehensive list, see the section on “list of organisms/infections to know” below.) The "smaller print" organisms that have come out before in Paper A include leishmaniasis and BK virus, which are both covered in this section Part 4 clinical syndromes is useful to put microbiology in context, and is also useful for the RCPA Part I/II vivas which I found more clinical than the written/practical.

Koneman https://www.amazon.sg/Konemans-Color-Textbook-Diagnostic-Microbiology/dp/1284322378/ Unfortunately, this edition is also getting a bit dated (2016). Extremely intimidating the first time I saw this book. Gets better once you read a bit through it and realise what are the important things you need to know (mostly at the start/end of chapters that talk about introductions to the various groups of organisms and microbiology techniques) and what are the less important stuff that is OK to read and forget. Definitely read through the introduction chapters as these are basics of microbiology! Chapter 3/4 on immunological and molecular methods - good to read through, but I preferred the chapters in MCM. Bacteriology - a lot of information, including a lot of organisms that you do not need to remember for exams (e.g. one of my favourite organisms I came across in this book was Staphylococcus condimenti which is found in soy sauce - but not really an organism you need to remember for exams!). It is however useful to know that the lists of endless corynebacteria and identification tables exist to be able to easily refer to them when you need to in daily practice. Mycology - this is less comprehensive than Larone, but I really liked reading through the section here as I found it gave me a very good introduction/overview to the topic. Essentially the fungi here are mostly "important" so it is good to know about what is mentioned here. Parasitology - same as mycology, I liked this section as it had a very good introduction/overview to the topic, and again the parasites here are mostly "important" Virology - afraid I can't comment on this as I don't remember much about the virology chapter here.

Larone's Medically Important Fungi https://www.amazon.sg/Larones-Medically-Important-Fungi-Identification/dp/1683674405/ Good textbook for mycology When reading through need to differentiate between important fungi and small print fungi Methods chapters are useful

Clinical Microbiology Procedures Handbook (I used the 4th edition, updated edition is https://www.amazon.sg/Clinical-Microbiology-Procedures-Handbook-Multi/dp/1683673980/) I read through sections of this book as I rotated through the labs (e.g. read the TB section when was in TB lab). Start and end of each section is particularly useful as gives introduction and interpretation information. Some parts of the 4th edition were a bit outdated by the time I read it - but there is a brand new 5th edition out now. I did read through quite a lot of this, and if time had permitted I would have liked to read through more of it before my Part I. Is also useful to be familiar with this as it's useful to easily reference it in practice.

Manual of clinical microbiology (I used the 11th edition, updated edition is https://www.amazon.sg/Manual-Clinical-Microbiology-Multi-Carroll/dp/1683674294/) I did not read the whole of this book! I used this book mostly for techniques (particularly liked the chapters on molecular and immunoassays) and as a reference when needed.

(Please note I am missing a good virology textbook here - this is because I already did a good amount of virology in my past and that together with other resources and guidelines was more than sufficient to get me through my exams. The virology textbook I used as a final year undergraduate >10 years ago was Flint's (updated version is https://www.amazon.sg/Principles-Virology-Multi-Jane-Flint/dp/1683670329/ which looks rather different from the version I used which was only a single volume) but this may be overly a basic science textbook. If you're not confident with basic virology it might be advisable to also find a good virology textbook.)

Websites and guidelines (Australian)

RCPA website

RCPA webinar series Watch all of these! Can be quite topical to current issues Quite a few topics covered in the series came out in my exams There are also a few webinars that are specifically for exam prep

Exam specific prep

Meet the examiner session from RCPA Path update – should be updated yearly https://www.rcpa.edu.au/Education/Disciplines/Microbiology/Exams/Examination-Advice

Past year exam questions

Optimise oral examination handbook (2022)

Additional quality management questions: https://www.rcpa.edu.au/Education/Laboratory-Management/Quality-Management/Docs/Quality-management-exam-questions-(1).aspx

Position statements, policies, guidelines I read most of these for Part I/II Some of these cover topics that came out in my exam Selective reporting of antimicrobials in Australia Modules - Ethics, Bullying/Harassment, Quality, etc Workshop slides Molecular Microbiology – the slides are uploaded every year, not too long after the workshop

Australia Department of Health and Aged Care website - www.health.gov.au

List of diseases that are notifiable or have vaccines https://www.health.gov.au/diseases

Links to PHLN laboratory case definitions and CDNA SONGS Communicable disease https://www.health.gov.au/health-topics/communicable-diseases

Can check for updates: Australian health documents https://www.health.gov.au/resources/publications Use filter: topics --> communicable diseases Can also use filters: Publication type --> case definition Publication type --> guideline CDNA guidelines https://www.health.gov.au/resources/collections/cdna-guidance-documents

State health departments Each state has information about specific diseases/infections that are of importance there, plus information about health alerts (links below under “list of organisms/infections to know”)

There are also some very nice guidelines on certain state websites

NSW department of health NSW disease control guidelines (a bit similar to the SoNGs, has things that are not in the SoNGs) https://www.health.nsw.gov.au/Infectious/controlguideline/Pages/default.aspx

Northern Territory department of health Quite a lot of specific disease guidelines including invasive group A streptococcus (and acute post strep glomerulonephritis), diphtheria, leprosy, malaria, NTM, syphilis)

Queensland department of health Really nice infection control guidelines Exposure to blood and body fluids Foodborne illness outbreak Healthcare worker vaccination MDRO Outbreak management

Perinatal infections Really important to know about perinatal infections and it came up quite a bit in my exams

ASID management of perinatal infections guidelines https://anzasid.sharepoint.com/sites/E-Knowledge/Shared%20Documents/Forms/AllItems.aspx?id=%2Fsites%2FE%2DKnowledge%2FShared%20Documents%2FANZPID%2FASID%20Management%20of%20Perinatal%20Infections%203rd%20Edition%2Epdf&parent=%2Fsites%2FE%2DKnowledge%2FShared%20Documents%2FANZPID&p=true&ga=1

Pregnancy care guidelines https://www.health.gov.au/resources/pregnancy-care-guidelines/table-of-contents Includes information on pregnancy and infections and screening https://www.health.gov.au/resources/pregnancy-care-guidelines/part-f-routine-maternal-health-tests/

ASHM - bloodborne viruses and sexually transmissible infections in antenatal care https://www.ashm.org.au/wp-content/uploads/2023/08/ASHM-BBVs-STIs-in-Antenatal-Care-Resource-2022.pdf

Immunisation

Immunisation handbook: https://immunisationhandbook.health.gov.au/vaccine-preventable-diseases

National immunisation programme schedule: https://www.health.gov.au/health-topics/immunisation/when-to-get-vaccinated/national-immunisation-program-schedule

More immunisation guidelines and info from NCIRS https://ncirs.org.au/health-professionals/ncirs-fact-sheets-faqs

National vaccine storage guidelines - Strive for 5 https://www.health.gov.au/resources/publications/national-vaccine-storage-guidelines-strive-for-5?language=en

National guidelines for yellow fever vaccination centres and providers (Australian Department of Health) https://www.health.gov.au/sites/default/files/2022-12/national-guidelines-for-yellow-fever-vaccination-centres-and-providers.pdf

NPAAC and NATA

May be useful to read through once before Part I. Definitely read through all the microbiology-relevant ones and know well before Part II

NPAAC standards https://www.safetyandquality.gov.au/our-work/accreditation/pathology-accreditation-standards NATA/RCPA accreditation https://nata.com.au/accreditation/medical-laboratory-accreditation-iso-15189/

HIV/HepB/HepC

Healthcare workers with blood borne viruses https://www.health.gov.au/resources/collections/cdna-national-guidelines-for-healthcare-workers-on-managing-bloodborne-viruses

Exposure prone and non-exposure prone procedures in Australia https://www.health.gov.au/sites/default/files/documents/2020/03/cdna-guidance-on-classification-of-exposure-prone-and-non-exposure-prone-procedures-in-australia-2017.pdf

NSW management of healthcare workers potentially exposed to HIV/HepB/HepC https://www1.health.nsw.gov.au/pds/Pages/doc.aspx?dn=PD2017_010

HIV

Australasian society for HIV, viral hepatitis and sexual health medicine (ASHM) and guidelines/resources: https://ashm.org.au/resources/

Testing Portal: https://testingportal.ashm.org.au

Antiretroviral guideline https://arv.ashm.org.au HIV care guide PrEP and PEP guidelines for HIV U=U guidance https://ashm.org.au/hiv/ https://www.ashm.org.au/hiv/hiv-management/ https://hivlegal.ashm.org.au/ (And there are probably many other guidelines!)

National HIV testing policy https://testingportal.ashm.org.au/files/ASHM_National%20HIVTestingPolicy_2020_HIV_.pdf

Hepatitis B

https://www.hepatitisb.org.au

https://www.gesa.org.au/public/13/files/Education%20%26%20Resources/Clinical%20Practice%20Resources/Hep%20B/HBV%20consensus%20Mar%202022%20Updated.pdf

National Hepatitis B testing policy https://testingportal.ashm.org.au/files/ASHM_TestingPolicy_2020_HepatitisB_07_2.pdf

Hepatitis C

https://www.hepcguidelines.org.au

National Hepatitis C testing policy https://testingportal.ashm.org.au/wp-content/uploads/2023/03/National_HepatitisC_Testing_Policy_v1-4_March_2023-v2.pdf

STI guidelines

Australian STI management guidelines for use in primary care:

https://sti.guidelines.org.au This has links to other guidelines

ASHM also has a lot of microsites https://contacttracing.ashm.org.au https://testingportal.ashm.org.au/ https://syphilisoutbreaktraining.com.au

Lifeblood (blood donation Australia) was an unexpectedly good resource for various infectious diseases

https://www.lifeblood.com.au/health-professionals/learn/resource-library

Filter for "factsheet" or search for "infectious" or search for "infections"

Should get a list of "transfusion focused infectious disease fact sheets" for a number of infections e.g. SARS-CoV-2, H5N1, Murray Valley Encephalitis Virus

These fact sheets are really nice as they summarise each infection including epidemiology in Australia, treatment, diagnosis, prevention Also useful for reading about transfusion associated infections!

Infection control and outbreak management

Australian guidelines for prevention and control of infection in healthcare https://www.safetyandquality.gov.au/sites/default/files/2023-09/australian_guidelines_for_the_prevention_and_control_of_infection_in_healthcare_current_version_v11.19_24_august_20232.pdf32.pdf

Outbreak management (Queensland guide) https://www.health.qld.gov.au/__data/assets/pdf_file/0025/444508/management-outbreaks.pdf

Infection control (Queensland) https://education.qld.gov.au/initiativesstrategies/Documents/infection-control-guideline.pdf

PHLN lab procedures and precautions for samples from patients with viral haemorrhagic fevers https://www.health.gov.au/resources/publications/phln-laboratory-procedures-and-precautions-for-samples-collected-from-patients-with-viral-haemorrhagic-fevers

Guidelines for legionella control https://www.health.gov.au/sites/default/files/documents/2022/07/enhealth-guidance-guidelines-for-legionella-control.pdf

Infectious diseases requiring additional precautions (NSW) https://aci.health.nsw.gov.au/__data/assets/pdf_file/0008/286838/ID_requiring_additional_control_precautions.pdf

GENCA/GESA guidelines on endoscope reprocessing https://www.genca.org/public/5/files/Nurses%20info/IPCE%202021_Feb2022update.pdf

RACGP infection prevention and control standards https://www.racgp.org.au/FSDEDEV/media/documents/Running%20a%20practice/Practice%20standards/Infection-prevention-and-control.pdf

Mycology

Mycology Adelaide https://www.adelaide.edu.au/mycology/ This was somewhat similar to Larone but is a good resource and is Australian

Additional useful guidelines and documents

Uptodate list of health hazards in travellers to Australia and NZ: https://www.uptodate.com/contents/potential-health-hazards-in-travelers-to-australia-new-zealand-and-the-southwestern-pacific-oceania

CARAlert organisms (critical antimicrobial resistances) https://www.safetyandquality.gov.au/our-work/antimicrobial-resistance/antimicrobial-use-and-resistance-australia-surveillance-system/national-alert-system-critical-antimicrobial-resistances-caralert

Australian Society for Microbiology - Guidelines for QC of microbiological culture media https://static1.squarespace.com/static/5c184125b98a781241e53ab6/t/5db2996892f5c03a7ff21a0f/1571985772520/Guidelines+for+the+Quality+Assurance+of+Medical+Microbiological+culture+media+2nd+edition+July+2012.pdf

Australian risk group organisms - I couldn't find a good resource for this! Some partial ones at https://cdn.csu.edu.au/__data/assets/pdf_file/0019/3283012/Biosafety-Manual-Booklet-3.pdf

Packaging and transport of infectious substances https://apps.who.int/iris/rest/bitstreams/1333766/retrieve#page17

Australian Refugee Health guide https://refugeehealthguide.org.au

List of Australian tests and diseases https://pathologytestsexplained.org.au

Infections disease transmission in solid organ transplantation https://tsanz.com.au/storage/documents/TSANZ-Infectious-Disease-review_Final_collated_edit.pdf

CDS https://cdstest.net – I found the resistance mechanisms explanations here very good

Websites and guidelines (non-Australian)

CLSI

M100 - I read this from cover to cover

Other CLSI documents that might be helpful include M35 abbrieviated identification of bacteria and yeast

M39 cumulative antibiogram

M45 ST of infrequently isolated or fastidious bacteria

M47 blood cultures - overlaps with the UK SMI and CMPH on blood cultures

M48 lab detection and ID of mycobacteria

M56 anaerobes

M58 MALDI

M62 ST of mycobacteria, Nocardia, other aerobic actinomycetes

MM03 molecular diagnostic methods for infectious diseases

MM09 nucleic acid sequencing methods

MM18 identification of bacteria and fungi by targeted DNA sequencing - basically 16S/ITS (And also other documents…)

EUCAST

Bacteria susceptibility testing document - useful to get some familiarity with this.

There are also a lot of helpful documents on the website that are good to read.

CDC

Infection control https://www.cdc.gov/infectioncontrol/index.html

CDC DPDx - lots of nice photos for parasites

CDC Yellow book https://wwwnc.cdc.gov/travel/yellowbook/2020/table-of-contents

Disinfection and sterilisation https://www.cdc.gov/infectioncontrol/guidelines/disinfection/index.html

CDC lab biosafety - biosafety in microbiological and biomedical laboratories (6th ed 2020)

Other useful resources

UK SMI

Cystic fibrosis lab guidelines https://www.cysticfibrosis.org.uk/sites/default/files/2020-12/Laboratory%20standards.pdf

European CDC website https://www.ecdc.europa.eu/en

IDSA guidelines https://www.idsociety.org/practice-guideline/alphabetical-guidelines/

APHL/ASM clinical lab preparedness and response guide https://www.aphl.org/aboutAPHL/publications/documents/WORK_BlueBook.pdf

Keeping up with current topics

Good to know about current topics in microbiology – this includes JCM etc for things like new methods, issues with testing, etc

Also look specifically for topics that are important in Australia Browse Australian news and see if there is anything that sounds important/likely E.g. Japanese encephalitis, mpox, flooding, black mould… If it has been reported in mainstream news and is related to infection it would be good to know about it!

National Notifiable Diseases Surveillance System (NNDSS) reports https://www.health.gov.au/resources/collections/nndss-fortnightly-reports - useful to get an idea of what is happening in Australia Each Australian state has their own site with health alerts – good to have a browse through to see if there is anything of particular interest.

NSW https://www.health.nsw.gov.au/Infectious/alerts/Pages/default.aspx

SA https://www.sahealth.sa.gov.au/wps/wcm/connect/public+content/sa+health+internet/public+health/alerts/health+alerts/

Queensland https://www.qld.gov.au/health/conditions/health-alerts WA https://www.health.wa.gov.au/Articles/F_I/Health-alerts-infectious-diseases

List of organisms/infections to know

When I first started microbiology training, the endless lists of organisms and infections seemed endless and extremely daunting.

I eventually settled on a "minimum" list of organisms/infections I need to know:

Anything that is mentioned in Part 3 of the Oxford Handbook of ID and Microbiology

Anything that is in the Australian guidelines (PHLN, CDNA SONGS, ACSQHC)

Anything that is in the Australian vaccination handbook

Anything that is listed in the infectious diseases lists of the individual Australian states

Victoria https://www.health.vic.gov.au/infectious-diseases/disease-information-and-advice

NSW https://www.health.nsw.gov.au/Infectious/factsheets/Pages/default.aspx

Queensland https://www.health.qld.gov.au/disease-control and https://www.qld.gov.au/health/condition/infections-and-parasites

NT https://health.nt.gov.au/public-health-notifiable-diseases

WA https://www.health.wa.gov.au/Articles/N_R/Notification-of-infectious-diseases-and-related-conditions

Alternative way of thinking about the organisms/infections are: Need to know well

Common and important, e.g. MSSA/MRSA, Enterobacterales, VZV, CMV

Common but unlikely (or less likely) to be clinically significant e.g. Staphylococcus epidermidis

Rare but important or with important associations, e.g. coccidioides, Mycobacterium ulcerans, Corynebacterium diphtheriae/ulcerans/pseudotuberculosis

Don’t really need to know well Rare and unlikely to be clinically significant e.g. Staphylococcus condimenti

For each organism/infection, I learned

Type/category of infection

Epidemiology – especially if there is anything specifically important to Australia, e.g. certain groups/regions/activities that have higher risk for infection

Transmission

Clinical presentation

Diagnosis

Non-laboratory diagnosis

Laboratory diagnosis

Sample type and collection

Sample transport

Microbiological diagnosis methods

Typing methods

Any particular issues with testing?

Treatment

Prevention

Anything special I needed to know

Specific additional things to look up for Part I

Not comprehensive!

Sensitivity, specificity, PPV, NPV - be able to calculate this - how this is affected by prevalence of disease

Receiver-operating characteristic curves

HEPA filters and how they work

Requirements of a PC2/3/4? laboratory

How is immunoglobulin avidity testing done (this is done for CMV in Australia?)

Sterilisation and disinfection

Types of autoclave and controls needed

Audit cycle

Outbreak management and epidemiological investigation

Cumulative antibiogram

Validation vs verification

Infection control - standard, contact, droplet, aerosol precautions

Measurement of uncertainty

R0 and R

Clinical trials and phases

PCR targets used for organisms

PK/PD

Australian infectious diseases screening programmes

Quality control and quality assurance

Levy-Jennings chart

Methods of diagnosis and specifics about how they work

Isothermal amplification

Sequencing by Sanger

Whole Genome Sequencing - Illumina, Nanopore

Different serological assays and how they work - ELISA, capture assays, haemagglutination-inhibition, CFT, virus neutralisation assays

Immunochromatographic tests

Transport and handling of infectious substances, how are they categorised (A/B)

Types of biosafety cabinets and how they work

Risk groups, definition of risk groups

Australian groups and organisations and regulations - NPAAC, NATA, TGA…

Organ/blood donation and microbiological testing

I also had definitions for

Quality control, quality assurance

Measurement uncertainty

MIC, ECOFF, breakpoints, susceptible, resistant, intermediate, SDD, wild type, non-WT

Critical concentration (MTB)

Sterilisation, disinfection, cleaning

R0, R

Validation, verification

Antibody titre

Universal and standard precautions

Prozone/hook effects

Notes about each part of the Part I exam

Disclaimer This is based on my own experience with the exam and there is always a possibility that the exam format or emphasis may change from year to year!

Paper A

RCPA publishes the previous papers for the past few years, and your seniors may have papers going back even longer.

Some of the questions repeat, so it is useful going through the past questions and preparing answers.

Keep in mind time limitations in the exam - there is a limit to how much you can write in 9 minutes! May be worth doing some of the questions "timed" to get a realistic idea of how much you can write. I think I wrote around a page for most questions.

A lot of basic microbiology here – e.g. “lab diagnosis of X” and technical questions.

Paper B

Short answer questions that cover a lot of various aspects of microbiology

Quite a few of the questions were "spot diagnosis" - photo provided, what is the likely organism, question related to this. Test ability to recognise fungi, parasites, etc.

Questions and topics were very random and were pulled from everything and everywhere.

Practical

Practice working up samples (including EQA samples) in the lab in parallel with routine work up done by lab staff

Make sure you are able to do all the tests that are known to be tested - e.g. I had to learn how to do urine microscopy as this isn't done by microbiology here. Double check what you need to be able to do for your year (my year included Gram stain, AFB stain, urine microscopy)

Go through past EQA reports

Go through lab manual and SOPs

A couple of months before I generated a list of media/reagents/etc I thought I might need to use during the practical. I went through this list with the lab to ensure that I would have these available for use when I need and “reserved” a stock of agar plates/etc for my use during the practical.

Before the practical I generated my own list of media/conditions to use for each sample type - and followed this list (with some additions depending on question stem) on day 1 of the exam to plate things out.

Viva

Practice questions with study group - we tried to do this similar to the exam when we would read the question, make short notes, and answer the question over zoom. Once done you might get additional questions to answer, such as in the viva.

I thought the Part I viva was more clinical and more guideline based than the rest of the Part I exam.

I found that knowing the Australian guidelines helped a lot with the viva.

Lactococcus garviae is a Gram positive coccus that somewhat resembles Enterococci. It is a known animal pathogen having originally been isolated from cases of bovine mastitis.  It also causes pneumonia in pigs, and haemorrhagic septicaemia in fish (Gibello, 2016).  Many recent publications in the literature relate to its impact on aquaculture where it is a major pathogen (Meyburgh, 2017).

I encountered my first clinical isolate of L. garviae (from the blood culture of a patient) in April 2012. A quick internet search found that L. garviae bacteraemia had been associated with raw fish consumption in Taiwan (Wang CY, 2007) and could lead to endocarditis. That of course captured my interest as I am a collector of exotic zoonotic pathogens. Our lab has been archiving any L. garviae we have isolated since then.

There have been 30 non-duplicate cases of L. garviae from 2012 to 2020. Most were isolated from blood (12), with the remainder from urine (6), wounds (4), bile (6), catheter (1), ear abscess (1). Often L. garviae was isolated with other bacteria in a mixed culture.

Of the wound isolates, three were from the finger, and in two cases where the mechanism of injury was documented, the patient had been chopping meat (not detailed).

Of the blood culture isolates, six were from post-mortem specimens and therefore of uncertain significance. Of the remaining six, three were from patients that underwent 2D-echo and had no evidence of endocarditis.  The remaining isolate was from a patient with a mitral valve bioprosthesis, who was managed for endocarditis even though no vegetations were seen on echo. Two patients did not have 2D-echo performed.

So L. garviae did not seem to be particularly associated with endocarditis in our local context but how were patients acquiring them? I knew from my studies on  Group B Streptococci that L. garviae are readily isolated from local fish.

In all our patients, there was no history of raw fish ingestion but taking an accurate food history in Singapore is a very fraught undertaking. There was also the possibility of other food sources given the animal reservoirs described above with L. garviae also being found in meat (Comi, 2020) and dairy (Scillieri, 2020) products.

With the help of 2 Ngee Ann Polytechnic students (Snow Lin and Kwek Kah Hao) we subjected all isolates in our archive collection from 2012-2018 to whole genome sequencing and compared them with whatever sequences of L. garviae that were available to see if we could find answers.

In total 11 local isolates were sequenced (Table).
 Virulence and antimicrobial resistance genes were identified. The strains were typed using multi-locus sequence typing (MLST) and compared with other published whole genome sequences from L. garvieae isolated overseas.

Local isolates had more resistance genes compared with the overseas isolates. 

Based on the MLST result, L. garviae can be divided into 3 lineages or genetic clusters (GC) (Figure). Non-human GC1 isolates overseas came from fish, meat and dairy sources.
 Non-human GC2 isolates overseas came from fish, plant and dairy sources.
 Most GC3 isolates overseas were from human sources (1 pork).
 We also noticed that some virulence genes (putative internalin and mucus adhesin) were present in GC2 and GC3, 
but not GC1. 


Local fish and human isolates were scattered across the 3GCs 
illustrating the heterogeneity of local isolates.
 

DB44798 from local human blood shared a common sequence type (ST) with isolate 138 from human urine in 
Spain and belonged to the same clonal complex (CC3) as FISH1 from local Cobia. Most CC3 
isolates overseas have so far been found in humans and dairy products.


DM12426 isolated from local human finger shared common ST41 with CT2 isolated from a local 
Bighead Carp.


Because the existing MLST database is not very discriminatory, we couldn’t really narrow down a specific food source for our human L garviae isolates, however it is also possible that they may really come from diverse sources.

This work has since been published (Lin YS, 2020).

 Gibello A, Galán-Sánchez F, Blanco MM, Rodríguez-Iglesias M, Domínguez L, Fernández-Garayzábal JF. The zoonotic potential of Lactococcus garvieae: An overview on microbiology, epidemiology, virulence factors and relationship with its presence in foods. Res Vet Sci. 2016 Dec;109:59-70.  (no free access)

Meyburgh CM, Bragg RR, Boucher CE. Lactococcus garvieae: an emerging bacterial pathogen of fish. Dis Aquat Organ. 2017 Feb 8;123(1):67-79.  (open access)

Wang CY, Shie HS, Chen SC, Huang JP, Hsieh IC, Wen MS, Lin FC, Wu D. Lactococcus garvieae infections in humans: possible association with aquaculture outbreaks. Int J Clin Pract. 2007 Jan;61(1):68-73. (no free access)

Comi G, Muzzin A, Corazzin M, Iacumin L. Lactic Acid Bacteria: Variability Due to Different Pork Breeds, Breeding Systems and Fermented Sausage Production Technology. Foods. 2020 Mar 13;9(3):338.  (open access)

Scillieri Smith JC, Moroni P, Santisteban CG, Rauch BJ, Ospina PA, Nydam DV. Distribution of Lactococcus spp. in New York State dairy farms and the association of somatic cell count resolution and bacteriological cure in clinical mastitis samples. J Dairy Sci. 2020 Feb;103(2):1785-1794. (open access)

On 31 Dec 2019, our associate consultant Dr Karrie Ko posted a Reuters report on the Department of Microbiology junior doctors WhatsApp chat- “Chinese officials investigate cause of pneumonia outbreak in Wuhan.” It didn’t elicit an immediate response. At that time most of us did not even know where Wuhan was, or that it had a population of over 11 million. It all seemed very remote.

My regular tea-buddy is from the Department of Molecular Pathology. Through Jan, the Wuhan pneumonia situation cropped up regularly in our discussions. If there was a need to diagnose this new disease by polymerase chain reaction (PCR), his department would bear the brunt of testing. Even back then, they were already making decisions on choice of swabs, and stockpiling reagents in anticipation of future shortages. They detected the first COVID-19 case in Singapore on 23 Jan2020. At that time I was just concluding a carefree holiday in Kyushu, Japan. My wife said there was a noticeable change in my mood when I got the news. I could tell from my WhatsApp chats and work emails that the hospital was mobilising as if for war.

On 6 Feb, my tea-buddy sent me an article reprinted in the Straits Times from Caixin Global titled "Reporter’s Notebook: Life and death in a Wuhan coronavirus ICU”. Up till then, I still harboured illusions that this could be a relatively mild illness like the H1N1 in 2009, in fact the initial cases did not seem to be as bad as SARS. However we now know that it is precisely because many people experience mild (or no) symptoms, that this virus is so successful at spreading. This article gave the first hint that if many people got infected, there would be enough very sick patients presenting in a short time to overwhelm a healthcare system.

Ramping up

Every Friday morning, the Singapore infectious diseases community meets at one of the larger public hospitals to discuss interesting cases. On 7 Feb, we met virtually on Zoom for the first time to discuss some of the earliest COVID-19 cases. We have not met in-person since.

The following day (Feb 8), the Infection Prevention Team required all staff to wear masks in the laboratory. This seemed strange as we were not patient-fronting, and we have been handling all specimens in biosafety cabinets to reduce risk of infection since SARS in 2003, . I half-joked that this was to protect us from infecting each other. I don’t know if this was indeed the original intention but in light of present universal masking guidelines, it probably was not a bad move.

The Department of Microbiology was largely spared the initial flurry of activity. However it soon became apparent that our molecular pathology colleagues were rapidly getting swamped with work. Immediately after Lunar New Year (28 Jan) we decided to deploy our microbiology trainees to help them. It was an easy decision to make. This was an once-in-a-lifetime (hopefully!) opportunity for the trainees to participate in a national effort during a pandemic of historic proportions. Whatever experience they acquired would still be very relevant to their training. So far, they have assisted in the evaluation of three automated SARS-CoV-2 PCR test systems, and the validation of pooled samples and different swabs. They also assisted in data collection and statistics, and reporting of SARS-CoV-2 results.

Dr Kenneth Goh (above) and Dr Shireen Tan (below) performing an evaluation of a SARS-CoV-2 PCR test.

Dr Deborah Lai (left) reviewing a SARS-CoV-2 PCR result in the Department of Molecular Pathology.

However as the national demand for testing surged, this proved to be still insufficient. From Feb 10 onwards, up to seven microbiology medical lab technologists/scientists (MLT/S) were reassigned to molecular pathology to help with the increasing load of SARS-CoV-2 PCR requests. This was a much more painful decision, as labs are only staffed just sufficiently to be cost-efficient during peacetime. This meant that we had to find ways of coping with less staff, and it was a scheduling nightmare for the supervisors. We even had to contemplate cutting services, though fortunately it never actually came to that.

Settling into a new routine

By April-May, the situation started to stabilize with molecular pathology having increased their staffing and capacity. We had also started introducing our own battery of COVID-19 tests.

In the Diagnostic Bacteriology Lab, MLT Yap Hock Guan loads a random-access automated PCR test for SARS-CoV-2 into a machine that is also used for MRSA and Carbapenemase-producing Enterobacterales screening. This particular test (introduced into service on 13 May) is used when a patient in the hospital needs an urgent result within a few hours. The Department of Molecular Pathology continues to do the vast majority of SARS-CoV-2 PCR testing in large batches.

Over in the Virology Lab, MLS Tan Chai Teng runs a high throughput COVID-19 serology test. To date, we have evaluated a total of eight different serology tests. We started offering this service from 14 May, and by the end of June we had run about 26,000 tests.

Under normal circumstances, the Epidemiology Laboratory carries out molecular fingerprinting of multidrug resistant bacteria for hospital outbreak investigations. Here MLT Jolene Gien loads a sequencer under the watchful eye of Senior MLS Dr Nurdyana Bte Abdul Rahman. They are sequencing SARS-CoV-2 genomes to help guide contact tracing efforts. These sequences (together with those contributed by other labs in Singapore and around the world) are uploaded onto the GISAID website (https://www.gisaid.org).

Dr Karrie Ko was starting her PhD on the hospital environmental microbiome when the COVID-19 outbreak occurred. She has now redirected some of her research to the study of SARS-CoV-2 in the environment.

It has been eight months (as of 1 Sept) since that initial WhatsApp post and we have settled into a kind of rhythm. We now feel naked without a mask, meetings are held by videoconference even if our offices may only be a short distance away from each other, meals are consumed in solitude on-site rather than with work-mates over at Tiong Bahru Food Centre. There is a SafeEntry QR code on my office noticeboard, and we have to log in our temperature recordings twice daily. I no longer meet my colleague from the Department of Molecular Pathology for tea.

Despite the stress and social isolation, everybody is taking things in the right spirit and is working with a common purpose. We are painfully aware of all the suffering around the world, and are grateful to be making a direct contribution to get Singapore back on track.

This is a longer version of an article first published in SMA News (reproduced with permission).

The first ever case of zoonotic fish-associated S. dysgalactiae subspecies dysgalactiae (SDSD) infection was isolated in our lab in 2005. This was during a period when we were actively looking out for cases of Streptococcus iniae, another fish-associated zoonosis (see an earlier post).

We had isolated a Streptococcus from the blood culture of a breast cancer mastectomy patient with cellulitis ascending her upper limb to the chest following a puncture injury of her index finger on the same side. She had been cleaning fish (Red Tilapia) and shrimps at the time. This is a typical setting for S. iniae infection. However, the isolate was alpha-haemolytic, PYR negative, and Lancefield group C. So this was not S. iniae which is beta-haemolytic, PYR positive, and not groupable, The API 20 STREP had identified it as S. dysgalactiae subs. dysgalactiae/equisimilis which is entirely in keeping with a large colony Group C Streptococcus. So nothing special here-or perhaps...?

After getting over my initial disappointment, I read that Streptococcus dysgalactiae (SD) comprise 2 distinct subspecies. The more commonly encountered Streptococcus dysgalactiae subspecies equisimilis (SDSE) are large colony β-hemolytic Group C and Group G streptococci that cause pharyngitis, skin and soft tissue infections in humans, much like  Group A Streptococcus pyogenes.

On the other hand, SDSD are classically described as α-hemolytic Group C Streptococci that cause disease in animals, especially cattle. And just recently, SDSD had also been found to cause disease in farmed fish...SDSD are not mentioned in textbooks of human clinical microbiology. So we were potentially sitting on the first case of zoonotic SDSD infection!

To cut a long story short we eventually were able to prove our isolate was indeed a fish associated SDSD by comparing pathogenicity gene sequences with those of fish isolates (Koh TH, 2009).

DB49998/05 the first case of zoonotic SDSD infection.

DB49998/05

We have since picked up a few other SDSD isolates.

DB60705/15

DB60705/15

DB60705/15 was an alpha-haemolytic Group G Streptococcus isolated in 2015 from blood culture of a patient with right breast cancer and lymphedema of the arm on the same side. This patient was discharged before we could interview her but Providencia stuartii, a known fish pathogen, was isolated from the same blood culture. Interestingly, this isolate also had the same multi-locus sequence analysis (MLSA) and emm type profile as DB4998/05.

STREP 97/15

STREP 97/15

STREP 97/15 was isolated from a sample of fish meat (Red Tilapia) purchased from a supermarket. This was during a large outbreak of Group B Streptococcus (GBS) in non-pregnant adults in Singapore (see https://10minus6cosm.tumblr.com/post/132590510556/serious-group-b-streptococcal-infection-in-adults), and we were investigating fish meat as a source of GBS. I was intrigued to find a beta-haemolytic (usually a sign of pathogenicity) Group G Streptococcus in the fish meat, and even more surprised to find it was SDSD (remember they are supposed to be alpha-haemolytic). However I subsequently found an article that showed that other researchers were also finding strains of SDSD that were beta-haemolytic (Jensen, 2012). Of course finding SDSD in a sample of fish meat does not prove that it was originally in the fish as we cannot exclude contamination by human handing of this fish. However when we did the molecular analysis, the pathogenicity gene profile, and alignment of gene sequences did indeed cause DB49998/05, DB60705/15, and STREP 97/15 to cluster with fish rather than human isolates.

DB53993/17

DB53993/17

DB53393/17 was a beta-haemolytic SDSD (non-groupable because of auto-agglutination) isolated in 2017 from a patient who had a mastectomy for breast cancer and presented with upper limb oedema and cellulitis on the same side. This time I had a chance to speak to the patient but try as I might I could not elicit a history of fish contact from her. She was a food stall owner who only handled chicken meat. As far as I could tell from my review of the literature till then, SDSD did not infect chickens. However when we did the molecular analysis, not only did DB53393/17 not cluster with human isolates, it was also closely related to an isolate that had been isolated from a chicken! In the late 1980s-early 1990s, there had been an investigation of focal dermatitis and cellulitis in broiler chickens in Canada that yielded beta-haemolytic SD. This article was easy to miss because SD were not explicitly mentioned in the title.

The genomes of these zoonotic SDSD have all been sequenced and it is clear that they form a separate cluster similar to animal isolates and distinct from  human isolates (Koh TH, 2019).

Clustering of human and animal isolates of SD based on tuf (elongation factor Tu) gene sequences. The length of the horizontal lines is in proportion to the differences between sequences (ie a longer horizontal line indicates sequences that are more dissimilar).

So summing up.

1. SDSD is a zoonotic pathogen. 

2. While it is likely to be rare, it is possible that SDSD is under-recognised in clinical microbiology labs as it can appear beta-haemolytic like the human pathogen SDSE. Phenotypic methods including MALDI-TOF are not good at discriminating SDSD from SDSE.

3. Women who have mastectomy and lymph node clearance for breast cancer are particularly susceptible to ascending infection and sepsis, and need to take care to avoid penetrating injuries when handling meat or fish.

Koh TH, Sng LH, Yuen SM, Thomas CK, Tan PL, Tan SH, Wong NS. Streptococcal cellulitis following preparation of fresh raw seafood. Zoonoses Public Health. 2009 May;56(4):206-8. (no free access)

Jensen A, Kilian M. Delineation of Streptococcus dysgalactiae, its subspecies, and its clinical and phylogenetic relationship to Streptococcus pyogenes. J Clin Microbiol. 2012 Jan;50(1):113-26. (free access)

Messier S, Quessy S, Robinson Y, Devriese LA, Hommez J, Fairbrother JM. Focal dermatitis and cellulitis in broiler chickens: bacteriological and pathological findings. Avian Dis. 1993 Jul-Sep;37(3):839-44. (no free access)

This story from my perspective begins on the 28 December 2015- at 5.27 pm precisely. I know because that exact moment is captured on the WhatsApp chat on my handphone.

That’s my virology colleague remarking that he was signing out more Hepatitis E results than he expected. I’m no virologist but it did seem odd to me too. Hep E virus (HEV) is a positive-sense, single-stranded, non-enveloped, RNA virus that is spread by the faecal-oral route. When I was a microbiology trainee I was taught it was a disease mainly of developing countries due to ingesting contaminated water and shellfish. The sanitation in Singapore is relatively good. Unless travellers were importing the infection, there was really no reason to find many cases of Hep E in Singapore. So why should there be a spike in cases when there really should hardly be any cases in the first place?

You can follow our train of thought below as we approached this conundrum.

The suggestion that it was zoonotic was a bit of a stab in the dark on the basis that HEV is acquired by ingestion. Given we have access to clean water in Singapore, a contaminated food source seemed a more likely explanation. We were also influenced by recent experience of a large Group B Streptococcus acquired by eating raw fish.

With some judicious googling, my colleague soon stumbled on a link between pigs and HEV which I had hitherto been unaware of.

Even if you find the virus in food, it still needs to be ingested in an infectious form. Aren’t people normally aware that you should only eat pork that has been thoroughly cooked (to avoid parasites)? Coincidentally, a few days before this conversation I had first hand experience of a potential risk exposure event (top photo). One of the local dishes in Singapore is congee where raw pigs liver is placed on top of hot rice porridge. As anyone who likes eating offal knows, there is nothing more dreadful than overcooked liver, so the connoisseur will try and time it such that the liver is barely just cooked. It was still bloody when I bit into it.

So we had a working hypothesis. Over the next few days a bit of digging around revealed a number of interesting facts.

1. When we looked up the Ministry of Health’s Weekly Infectious Disease Bulletin, we found that there were more cases of Hep E (97) than Hep A (70) reported in 2015.

2. A lot of other developed countries were noticing unexpected increases of Hep E cases and these were increasingly linked to undercooked pork products (particularly sausages and pates).

3. Hep E had been already been studied in Singapore in 2013 (Tan LTC et al, 2013). At that time the association with pigs had in fact been considered but there was insufficient evidence to draw a firm conclusion about the food source, partly because the molecular tools to genotype HEV were not widely available at the time. HEV genotypes 1 and 2 are restricted to humans and cause the traditional outbreaks found in developing countries as a result of poor sanitation. Genotypes 3 and 4 may be found in humans but also have animal reservoirs, in particular pigs. 

So we embarked on a new study together with colleagues in the National Environmental Agency (now Singapore Food Agency) which has finally been published (Wong CC, 2019).

The findings can be summarised thus.

1. Hep E has been increasing in Singapore. From 0.92 cases per 100,000 population in 2009-2011 to 4.1 cases per 100,000 population in 2016.

2. The increase is predominantly in the Chinese population. Chinese (86%) were over-represented among those HEV IgM positive compared to Indians (5%) and Malays (2%), even taking into account the National ethnic distribution.

3. The majority (85%) of HEV were genotype 3. 15% were genotype 1 and 1 was genotype 4. Genotype 3 was found in both pig livers and human infections among the Chinese.

4. A different genotype of HEV which is not associated with animal sources was found in the Malays (unsurprising since Muslims do not eat pork).

During the long gestation time for this paper, a number of other local studies have been published.

A fatal case of chronic Hep E in  a solid organ transplant recipient (SOTR) in our hospital triggered a retrospective review of 16 cases, eight of which were SOTR (Teo EC et al, 2017)*. 88% were HEV genotype 3. 

Earlier this year another team of Singapore investigators  (Zhu YO, 2019) published on viral sequences from 15 immunocompromised patients with chronic Hep E at the National University Hospital. All were HEV genotype 3. Interestingly they found two patients who shared extremely similar viruses who had residential addresses, which were within one kilometre of each other suggesting  the possibility of common food sources.

So the conclusion is that eating raw or undercooked pork products may contribute to the majority of Hep E cases in Singapore.

What are the risks?

In most healthy people, Hep E is a mild infection and may not even be noticed.

However it may cause chronic infection in those with pre-existing liver disease and more severe infection in the immunocompromised.

There are implications here for public health and food for thought for those looking after immunocompromised patients and those running blood banks.

Preventative food safety precautions may be found in some of the links given below.

UK Food Standards.

https://www.food.gov.uk/safety-hygiene/hepatitis-e-virus

Center for Food Safety Hong Kong 

https://www.cfs.gov.hk/english/programme/programme_rafs/programme_rafs_fm_01_15_HEV_pig_liver.html

https://www.cfs.gov.hk/english/multimedia/multimedia_pub/multimedia_pub_fsf_54_01.html

European Food Safety Authority

https://efsa.onlinelibrary.wiley.com/doi/epdf/10.2903/j.efsa.2017.4886

Tan LT, Tan J, Ang LW, Chan KP, Chiew KT, Cutter J, Chew SK, Goh KT. Epidemiology of acute hepatitis E in Singapore. J Infect. 2013 May;66(5):453-9. (no free access)

Wong CC, Then SM, Ng Y, Kang JSL, Ng TY, Chau ML, Koh TH, Chan KP. Seroepidemiology and genotyping of hepatitis E virus in Singapore reveal rise in number of cases and similarity of human strains to those detected in pig livers. Zoonoses and Public Health. 2019 (early view online). (no free access)

Teo EC, Tan BH, Purdy MA, Wong PS, Ting PJ, Chang PJ, Oon LL, Sue A, Teo CG, Tan CK. Hepatitis E in Singapore: A Case-Series and Viral Phylodynamics Study. Am J Trop Med Hyg. 2017 Apr;96(4):922-928. (free access)

Zhu YO, Aw P, Aung MM, Lee HK, Hibberd M, Lee GH. Patterns of mutation within an emerging endemic lineage of HEV-3a. J Viral Hepat. 2019 Jan;26(1):191-198. (free access)

Let’s say hypothetically one day you come across this Gram stained slide  of a positive blood culture from a patient on the haematology unit.

Yup, it is a mould. Those are hyphae. We don’t often see moulds in blood cultures but there is one that typically infects haematology patients. Fusarium spp. OK, these are immunocompromised patients who are more susceptible to fungal infections, fungi are everywhere, especially in the tropics, bad things happen.

Over the next 3 weeks you get another 2 cases of Fusarium fungaemia. All 3 patients are on the same unit. You check that they were not admitted with the infection-they weren’t. Now this is a bit more disconcerting. Intuitively you know that this is more than what you would expect. However to be sure you extract data for all Fusarium spp. isolated from blood cultures  in the last 8 years as recorded in the Laboratory Information System. Up to the previous month there have been only 3 additional cases, and none in the last 3 years.

Time to contact the infection control team. What is the likely source? Fusarium spp. are readily found in the environment (has there been any renovation work performed around the ward? Was there a contaminated product in common use?), but are particularly a pathogen of plants (has someone been bringing cut flowers into the unit?-surely not). But we also know from experience that Fusarium spp. are also readily found in the wet environment. A large outbreak of F. solani complex keratitis in 2005-2006 was thought to be due to contact lens solution bottles being contaminated in the patient’s home bathroom environment.

In the meantime you do a Pubmed search for outbreaks of Fusarium spp. 

There aren’t many!

In Greece, a Fusarium verticillioides outbreak among immuncompetent patients was associated with reconstruction works. Environmental samples were negative, though water sources were not very thoroughly investigated. (Georgiadou SP, 2014).

There have been a number of small defined outbreaks described in Brazilian haem-oncology units. One (Litvinov N, 2015) found invasive F. soloni and F. oxysporon associated with positive cultures in the taps, showers, drains and room air, whereas another (Carlesse F et al, 2017) could find no environmental source.

A much earlier study from a Texas Medical Center (Anaissie EJ, 2001) had in fact already investigated the risk posed by the hospital water distribution system, though this was not in the context of a defined outbreak. They noticed that there were molecular matches between patient and environmental isolates, and concluded that the water distribution system of a hospital could be a reservoir for Fusarium spp. (and other pathogens).

Fusarium species was recovered from 57% of water system samples. 88% of sink drains grew F. solani; 16% of sink faucet aerators and 8% of shower heads yielded F. oxysporum. F. solani was isolated from the hospital water tank. Aerosolization of Fusarium species was documented after running the showers. 

On the basis of their findings, the authors recommended that hospitals with cases of fusariosis (not of community-onset) should consider testing their water supply for the presence of opportunistic moulds. If such fungi were recovered from the water system, policies to avoid or minimize exposure of immunosuppressed patients to tap water from any source should be implemented (e.g. by providing boiled water for drinking). In addition, patients should avoid showering during severe immunosuppression because of the risk of acquiring the organisms through aerosolization of contaminated water. They recommend that bed baths with sterile disposable sponges be performed instead.

So is it in fact possible to isolate Fusarium spp from a shower head?

Can’t actually detach the shower head plate in this case, so sampling can only be performed where the head is detached from the hose at the handle. 

Looks fairly clean as you would expect. Just water, no gunk or visible biofilm.

Oh well...

Acknowledgment: Thanks to the Department of Infection Prevention and Control, SGH for assistance with the shower photos.

Georgiadou SP, Velegraki A, Arabatzis M, Neonakis I, Chatzipanagiotou S,Dalekos GN, Petinaki E. Cluster of Fusarium verticillioides bloodstreaminfections among immunocompetent patients in an internal medicine department after reconstruction works in Larissa, Central Greece. J Hosp Infect. 2014 Apr;86(4):267-71. doi: 10.1016/j.jhin.2014.01.011. Epub 2014 Feb 27. PubMed PMID: 24650721. (no free access)

Litvinov N, da Silva MT, van der Heijden IM, Graça MG, Marques de Oliveira L, Fu L, Giudice M, Zilda de Aquino M, Odone-Filho V, Marques HH, Costa SF, Levin AS. An outbreak of invasive fusariosis in a children's cancer hospital. Clin Microbiol Infect. 2015 Mar;21(3):268.e1-7. doi: 10.1016/j.cmi.2014.09.004. Epub 2014 Oct 12. PubMed PMID: 25658562. (free access)

Carlesse F, Amaral AC, Gonçalves SS, Xafranski H, Lee MM, Zecchin V, Petrilli AS, Al-Hatmi AM, Hagen F, Meis JF, Colombo AL. Outbreak of Fusarium oxysporum infections in children with cancer: an experience with 7 episodes of catheter-related fungemia. Antimicrob Resist Infect Control. 2017 Sep 7;6:93. doi: 10.1186/s13756-017-0247-3. eCollection 2017. PubMed PMID: 28912948; PubMed Central PMCID: PMC5588724. (free access)

There is general advice and exam specific advice (highlighted text has hyperlinks).

General Advice

This applies to preparing for local practice in Singapore. This has a slightly American slant (unfortunately the exams you will be sitting are British and Australian).

Refer to your JCST Seamless Programme Medical Microbiology Training Syllabus and use this as a checklist. Literally tick off each item as you go so that there are no gaps in your training experience.

During your lab training you will be used to seeing tests as individual items. However you must get your head round to seeing things also from the clinician’s perspective. For a patient presenting with a particular syndrome which battery of tests are appropriate for that individual patient? Even for a single pathogen (particularly with viruses), there may be a choice of different tests that can be used. Which are most appropriate for that particular situation? This is one of the characteristics that distinguishes medical student knowledge from that of a microbiology trainee. 

I suggest right at the beginning of training to get the fundemental cliinical perspective in place by reading Infectious Diseases A Clinical Approach by Yung et al. This is book is simple enough to be read in a fortnight. (Update Jan 2020: The new kid on the block is Comprehensive Review of Infectious Diseases by Spec A et al-which comes with an ebook with over 400 nicely written MCQs-unfortunately it probably can’t be finished in 2 weeks).

Please get yourself a microbiology textbook and systematically read it from cover-to-cover. The most appropriate at time of writing is the latest edition of Koneman’s Color Atlas and Text book of Diagnostic Microbiology. You should try to complete reading this book within your first year of training.

This will not be enough for Virology. Some may find Khare Guide to Clinical and Diagnostic Virology too simple but I think reading it cover-to-cover is still useful for the 1st year trainee. For subsequent years you will need to read selectively  Loeffelholz Clinical Virology Manual for the lab perspective and maybe dip into Richman Clinical Virology for the clinical approach.

The following websites are useful

For parasitology

CDC DPDx

Gorgas Course Case of the Week

CREEPY DREADFUL WONDERFUL PARASITES

For mycology

Mycology Online

One of the first things you should acquaint yourself with is the local lab, and hospital, biosafety and disinfection/sterilization policies. For lab biosafety the freely available BMBL should suffice as a reference.

In practice, you can expect to give a lot of advice on specimen collection so please familiarize yourself with our Pathology Handbook (and know where to find it). For reference you should look at A Guide to Specimen Management in Clinical Microbiology.

You should also familiarize yourself with our lab manuals (and those of the labs you visit when posted out-ask yourself why they are doing things differently from us).

For technical details you should also refer to the Clinical Microbiology Procedures Handbook.

There are 2 books you will always be consulting as references

Mandell, Douglas, and Benett’s Principles and Practice of Infectious Diseases Use this when you are following up interesting cases or building up your case portfolio

ASM Manual of Clinical Microbiology Not to be used as your primary textbook!

The most useful journals to read are the Journal of Clinical Microbiology, Clinical Microbiology Newsletter and Clinical Microbiology Reviews.

Please also look at the Cumitech Collection and the CLSI documents.

For infection control I like Lautenbach Practical Healthcare Epidemiology. For more details you should be keeping up to date with journals like Infection Control and Epidemiology and the Journal of Hospital Infection.

By the time you start preparing for the Part 2 exams you should be dealing with, and reading up on, management issues.

You should have helped trouble shooting of QA failures, and be familiar with the other CLSI documents that are relevant to lab practice but not microbiology specific. You should have been involved in at least one CAP accreditation inspection and should have passed the CAP inspector course itself.

The most useful lab management textbooks are

Garcia Clinical Lab Management Wagar Lab Administration for Pathologists Varnadoe Medical Laboratory Management and Supervision

But you should also familarize yourself with the local regulations governing labs and your hospital policy for issues like staff discipline etc…

Royal College of Pathologists (RCPath)

Look up the FRCPath training syllabus and tick off items to make sure you have covered all the ground. Look up past-year exams so that during your training you get a sense which lab situations you encounter are likely to come up as exam topics.

The principle of the postgraduate exams is they test you for your suitability to be a consultant. Because local practice is more lab-based, you will have to simulate the UK training environment.

A UK trainee may handle 5-10 positive blood cultures a day followed by a ward round that covers another 5 patients with infection issues like osteomyelitis or septic arthritis, AND a daily ICU round. They may encounter an infection control scenario every 1-2 weeks (needlestick, outbreak, staff exposure etc…). For this, they are giving specific advice including further investigations, antibiotic choice, dosage (remember they are also advising on therapeutic drug moniitoring for vancomycin and aminoglycosides), infection control measures, etc…

Even though we do not manage patients like they do in the UK you should build up your own personal case portfolio based on what you report in the lab. Do not stick to the minimum requirement in the syllabus! Actively follow up these cases and imagine how you would manage these patients if you were the doctor-in-charge.

Get the Oxford Handbook of Infectious Diseases and Microbiology, read it cover-to-cover and use it to guide your ‘management’. How would you adjust the dose for renal imparment? What alternative would you use if there was penicillin allergy?

UK trainees often refer to Kucer’s The Use of Antibiotics when finding out details about antibiotics so you should read it up for at least the common antibiotics that are commonly used (in Singapore and the UK).

Read through the editorials and guidelines in the Journal of Antimicrobial Chemotherapy going back at least 2 years (esp guidelines and new antibiotics). Check out the British Society for Antimicrobial Chemotherapy website.

Two books I find useful for selective reading are Fungal Infection: Diagnosis and Management by Richardson and Warnock. Lecture Notes on Tropical Medicine by Beeching and Gill

I have not got a copy of the Communicable Disease Control and Health Protection Handbook yet but it looks like it could be handy (Update Jan 2020: I have a copy now and it is useful for RCPath trainees).

You should also construct scenarios for infection control and outbreak management to supplement your actual experience. A lot of exposure management (e.g. VZV, measles, hepatitis B) can be found in the Green Book.

Go through the IPC in 5 videos. This is part of the trainee education program. Every UK trainee knows this material. So you should too.

Read through the editorials and guidelines in the Journal of Hospital Infection going back at least 2 years. Check out the Hospital Infection Society website.

You should subscribe (or have access ) to the UKNEQAS Microbiology Interpretive Comments.

You will also need to pretend that you are a Consultant Microbiologist practicing in the UK. Unlike the US, there is a lot more central control in the UK so all UK consultants receive and are expected to comply with the same set of guidance documents.

I could create my own list of useful websites where such documents may be found but microregistrar has already done it for me. PHE, HTM (e.g. dealing with water  supply in hospitals)  are particularly relevant. I don’t know if questions ever came out from the DH Building Notes, but it may be useful to know what relevant documents are in there (like HBN 00-09).

Don’t forget to scour the RCPath website and even read the bulletin going back 2 years for any material that may be relevant (like audit reports, documents or discussion of current college issues).

And don’t forget the UK follows ISO15189 for lab accreditation so you probably should know a bit about UKAS (How would you perform measurement of uncertainty for a PCR test?).

In my opinion, the FRCPath is quite a straightforward exam, and paradoxically quite a difficult exam to pass at the same time. They are not likely to ask anything a District General Hospital Microbiologist will not encounter, so no exotic bugs here. If you are fully involved in daily lab activities you may even be better prepared to sit the exam purely on accumulated experience rather than mugging standard textbooks. But as you can see from this list-if you do not know what to spend your energy on, fail to ensure you accumulate appropriate experience, or if you do not read the right material you could end up with close to zero exam-specific knowledge.

(Update Jan 2020: Infectious Diseases, Microbiology and Virology: A Q&A Approach for Specialist Medical Trainees is a new MCQ book which is useful for RCPath candidates. I am not that keen on the format of the MCQs (prefer Spec et al) but the selection of topics will give you some idea of the domain knowledge college examiners are looking for).

Royal College of Pathologists of Australasia (RCPA)

The RCPA seems to have changed to be more similar to the RCPath in the sense that they are now testing more practical consultant competence and less esoteric knowledge.

Therefore go through the same process as for the FRCPath.

Investigate the RCPA website for the training syllabus and other issues that Australian pathologists are currently grappling with.  For Part 2, relevant issues may not be obviously microbiological (like how do you handle bullying in the lab?). The RCPA actually put a lot of training material onsite including the National Microbiology Webinar series (need to be RCPA registered trainee for access). It goes without saying you should go through all of them.

Be aware they have their own Antibiotic Guidelines Book.

Other Australia specific websites (equivalent to those mentioned for RCPath) are

Australian Committee on Safety and Quality in Health Care

Department of Health

NATA

NHMRC

Australian Immunisation Handbook

Australian STI Guidelines

SA Health Antimicrobial Guidelines

ASHM testing portal

And like their marsupials, Australian microbiology has its own distinctive flavour. e.g. Legionella longbeachae, Cryptococcus gattii, Mycobacterium ulcerans, Hendra virus etc... So be familiar with what’s happening Down Under.

Hmmm...this turned out much longer than I expected. Hope it won’t scare off any prospective trainees...

We have previously blogged about using the Olympus Air for taking images of Gram stains. This gave the image quality of a dedicated microscope camera but the added convenience of quickly sharing the image because it was transferred to a handphone. But why not just use a handphone to capture the image directly?

Anyone who has tried this will realise it is not as easy as it sounds. If you just hold the handphone lens flush against the microscope eyepiece, it is very difficult to get a good image of the slide because the wide angle of the camera lens tends to include the sides of the eyepiece barrel. You can obtain a better field of view of the slide by holding the handphone a small distance away from the eyepiece but then you need steady hands...

We have experimented with a number of commercial microscope-handphone adapters without much joy. They all suffer from 2 problems. Firstly, they all hold the handphone flush with the eyepiece so you don’t solve the field of view problem mentioned above. Secondly, the attachment method does not result in a stable platform.

We were so frustrated with the results that we considered designing and 3D-printing our own adapter. It would have set the handphone lens a bit further back from the eyepiece and would involve a redesign of the attachment method that would leverage somehow on the binocular eyepiece of the microscope to keep the whole platform steady.

Someone has now come up with a better design based on a similar thought process.

The LabCam addressed the field of view issue optically by including a lens which replaces the microscope eyepiece (2 are provided which should fit most microscopes). Both the replacement eyepiece and the design of the adapter itself ensure a stable platform. It is easy to set up and use.

The LabCam was actually designed by an undergraduate student and converted to viable product by a Bench to Bedside Initiative.

There remain a number of issues.

Because it includes lenses, it is more expensive than the other adapters (but the ease of use and the results obtained will trump price in our view).

The lens could be of higher quality. Sharpness is good centrally but falls away at the edges. There also appears to be chromatic aberration at the edges (blue fringing around the image).

It is only available for iPhone at the moment, though the modular design does suggest the potential to apply the same concept to other handphones.

Nevertheless, we feel this is the best solution on the market at the moment. You can see more images captured with this system on our Instagram.

The SGH laboratory has been doing antimicrobial surveillance testing on regular  systematic sample collections of Neisseria gonorrhoeae sent to us from the Department of  Sexually Transmitted Infection Control, National Skin Centre since 1992 (at least). These results are submitted to the World Health Organization Gonococcal Antimicrobial Surveillance Programme (WHO GASP). If you are interested, the reports are freely available at the WHO GASP website (https://www.who.int/reproductivehealth/topics/rtis/gonococcal_resistance/en/).

Local isolates of N. gonorrhoeae are often resistant to penicillin, tetracycline, and ciprofloxacin. The key antibiotic is therefore a third generation cephalosporin like ceftriaxone, with azithromycin thrown in for good measure (https://www.dsc-clinic.sg/Healthcare-Professionals/Books%20and%20Publications/Pages/STI-Management-Guideline-6th-Edition-(2013).aspx).

Ceftriaxone has remained a reliable antibiotic for a surprisingly long time but resistance has finally emerged. The first ceftriaxone-resistant N. gonorrhoeae was reported in Japan in 2011 (Ohnishi M, 2011). This strain named H041 had a ceftriaxone MIC of 2 mg/L. Since then, ceftriaxone-resistant N. gonorrhoeae have been reported in an increasing number of countries. Resistance is due to a mosaic penA allele (i.e. the gene is made up of bits of DNA from different genomes, N. gonorrhoeae can do this because they can acquire DNA from their environment by a process called transformation). This results in an altered penicillin-binding protein with reduced affinity to beta-lactam antibiotics.

A particular clone, named after the original strain FC428 isolated in Japan in 2015 has spread internationally (Lee K, 2019). In 2017, the World Health Organization published a list of bacteria for which new antibiotics are urgently needed. Cephalosporin-resistant N. gonorrhoeae were listed as Priority 2: High.

Our lab diagnosed the first ceftriaxone-resistant N. gonorrhoeae to be isolated in Singapore in early 2018.

The isolate was resistant to ceftriaxone with an MIC of 1 mg/L (left) and an annular radius of 5 mm by the CDS method.

Whole genome sequencing was performed and analysis showed that the strain had the same penA allele as the FC428 clone. Furthermore it belonged to ST13871 (allele profile abcZ 126 adk 39 aro E67 fumC 987 gdh 148 pdhC 153 pgm 65) which is a single locus variant of ST1903 (allele profile abcZ 126 adk 39 aroE 67 fumC 157 gdh 148 pdhC 153 pgm 65), the common multi locus sequence type of the FC428 clone. More details can be found in our paper which has just been published (Ko K, 2019).

As many of the cases of ceftriaxone-resistant N. gonorrhoeae were reported to have been acquired in south-east asia, we can only expect this problem to become worse.

Ohnishi M, Saika T, Hoshina S, Iwasaku K, Nakayama S, Watanabe H, Kitawaki J. Ceftriaxone-resistant Neisseria gonorrhoeae, Japan. Emerg Infect Dis. 2011 Jan;17(1):148-9. doi: 10.3201/eid1701.100397. PubMed PMID: 21192886; PubMed Central PMCID: PMC3204624. (free access)

Lee K, Nakayama SI, Osawa K, Yoshida H, Arakawa S, Furubayashi KI, Kameoka H, Shimuta K, Kawahata T, Unemo M, Ohnishi M. Clonal expansion and spread of the ceftriaxone-resistant Neisseria gonorrhoeae strain FC428, identified in Japan in 2015, and closely related isolates. J Antimicrob Chemother. 2019 Apr 19. pii: dkz129. doi: 10.1093/jac/dkz129. [Epub ahead of print] PubMed PMID: 31002306. (no free access)

It seems that every month there is a new article about a carbapenemase-producing Enterobacteriaceae (CPE) outbreak traced to sinks (please read about some local experience here and here). There is now even a sink laboratory! However there may be other reservoirs to consider in the hospital wet environment.

We recently were trying to trace the environmental source of a clone of CPE. Environmental swabs of the usual suspects-sinks, toilet bowl, toilet drains were all negative. 

One tends to think of the sink and toilet first as potential reservoirs of CPE because patient secretions naturally find their way there. In retrospect, the shower drain is no less likely a place (especially if the patient is colonised or infected with CPE on the skin). In this case, the shower drain was the only environmental site sampled that was positive for CPE.

Perhaps shower drains may become the ‘next big thing’ when it comes to investigating CPE outbreaks.

Around the same time we were carrying out our investigation, an Australian team (Fernando SA et al, 2018) published a study (in a very similar clinical setting) where CPE was found to be heavily concentrated in the floor drains of the shower rooms (though that is not obvious from the title the paper). What is interesting about this article is that there was increased detection of CPE in the shower floor drains after cleaning as compared to before cleaning! This is similar to our own experience. 

Now if you were to look down a typical shower drain you can see why. There is a lot of biofilm down there! This is not easy to remove completely and the cleaning process itself disrupts the biofilm resulting in greater release of CPE.

But isn’t the floor drain downstream of the patient? Is it just a case of a patient with CPE contaminating the floor drain? Like the situation with the sinks, we still need to think of a mechanism whereby CPE in the drain can somehow transfer to another hitherto uncolonised/uninfected patient.

A recent study has shown that using the shower produced aerosols containing carbapenemase-producing Pseudomonas aeruginosa from the floor drain (Hopman J et al, 2019). 

In the long term this may require an engineering solution. A German group trying to control an outbreak of multidrug-resistant P. aeruginosa in a paediatric haematology unit redesigned their shower drains to be easily cleaned and disinfected and to reduce splashing (Kossow A, 2017).

Fernando SA, Phan T, Parker C, Cai T, Gottlieb T. Increased detection of carbapenemase-producing Enterobacteriaceae on post-clean sampling of a burns unit's wet surfaces. J Hosp Infect. 2019 Feb;101(2):179-182. (no free access)

Hopman J, Meijer C, Kenters N, Coolen JPM, Ghamati MR, Mehtar S, van Crevel R, Morshuis WJ, Verhagen AFTM, van den Heuvel MM, Voss A, Wertheim HFL. Risk Assessment After a Severe Hospital-Acquired Infection Associated With Carbapenemase-Producing Pseudomonas aeruginosa. JAMA Netw Open. 2019 Feb 1;2(2):e187665. (free access)

We have previously posted the case of a Streptococcus pneumoniae masquerading as Neisseria meningitides (see earlier post Streptococcus pneumoniae or Neisseria meningitidis?) We now present the converse situation.

N. meningitidis is a Gram-negative diplococcus, The bacteria in these images don’t really demonstrate the kidney- or coffee- bean shape described in textbooks. The main clue that they are Neisseria spp. are the fact that the cocci occur in pairs.

Image 1.

Image 2.

Image 3a 

Image 3b

Image 4a

Image 4b

Note that N. meningitidis sometimes resist the tendency to decolonize and may appear Gram-positive (Images 5 and 6).

Image 5a

Image 5b With ‘the eye of faith’ it is possible to discern a capsule (clearing of background material) around this apparently ‘Gram-positive’ diplococcus.

Image 6a

Image 6b There is a suggestion of capsule around these as well

Streptococcus minor is a recently described Gram positive coccus found in the tonsils and gastrointestinal tract of dogs, cats and a calf (Vancanneyt M, 2004). The most closely related species by 16S rRNA gene sequence similarity are the animal Streptococci-S. ovis (95.9%), S. gallinaceus (95.5%), S. iniae (95.4%) S. entericus (95%), and S. suis (94.1%).

Small colonies (hence minor) on sheep blood agar are surrounded by a narrow zone of alpha-haemolysis.

Gram stain of colonies growing on sheep blood agar.

Gram stain of colonies growing in broth culture.

The first case of human infection described was that of a 62 year old Dutch patient with peritoneal dialysis associated peritonitis (Dresselaars HF, 2014). The same strain of S. minor was isolated from the dialyse and the saliva of the patient’s cat.

The second case was a 51 year old Belgian woman who was bitten by a dog (Tré-Hardy M, 2016)

We did a retrospective review of all S. minor isolated in our laboratory from 2013 (when we started using MALDI-TOF for identification of bacteria) to 2017. The first isolate was in 2015 and there were a further 5 isolates from non-duplicate patients during the study period (3 each in 2015 and 2017). All patients had soft tissue infections involving pet bites (4 dogs and 2 cats).

This work was presented at the 28th European Congress of Clinical Microbiology and Infectious Diseases 2018. You can download and view the poster at the link below.

https://f1000research.com/posters/8-26

Vancanneyt M, Devriese LA, De Graef EM, Baele M, Lefebvre K, Snauwaert C, Vandamme P, Swings J, Haesebrouck F. Streptococcus minor sp. nov., from faecal samples and tonsils of domestic animals. Int J Syst Evol Microbiol. 2004 Mar;54(Pt 2):449-52. (free access)

Dresselaars HF, Zwart B, Pettersson AM, Rijnsburger MC, Ho-dac-Pannekeet MM. Peritoneal dialysis-associated peritonitis of zoonotic origin, when minor gets major. Neth J Med. 2014 Dec;72(10):551-3. (free access)

We are beginning an occasional series of blog posts on virology topics from our virology consultant Dr Wan Wei Yee.

Unfortunately Virology topics make for lousy blogging material as there are fewer opportunities for colorful and interesting pictures, but there is still scope for interesting discussions J. Apologies in advance for the wordy post…

Let’s discuss the investigation of OBI as I’ve observed many incomplete or inappropriate tests ordered prior to immunosuppressive treatment.

OBI is characterized by the persistence of Hepatitis B virus (HBV) genome in the liver of hepatitis B surface antigen (HBsAg) negative individuals, with/without detectable low level viraemia and with/without seropositive markers (e.g. anti-HBcore).

After acute HBV infection, the disappearance of HBsAg, presence of Hepatitis B core antibody (anti-HBcore)  +/-  Hepatitis B surface antibody (anti-HBs) indicates resolution. However, even after resolution, the virus lies dormant in the hepatocyte due to its ability to form a viral mini chromosome (cccDNA) within the hepatocyte nucleus which is kept in a latent state by the host’s immune system. That is until an opportunity arises leading to reactivation.

There is currently no curative and eradicative therapy available. Therefore, the purpose of screening for HBV prior to immunosuppressive treatment is to identify those who had previous exposure to HBV so that measures can be taken to prevent reactivation.

Why is this important? The prevalence of HBV infection in Singapore is intermediately endemic (5-6% in the 1980’s). Even though the incidence has decreased (3.6% in the 2010 seroprevalence study) as a result of the introduction of HBV vaccination in the childhood immunization program since 1987, nearly a quarter of the population have been exposed to HBV (22.5% anti-HBcore positive).1 Thus, OBI remains important in this substantial cohort due to it’s potential for reactivation as a result of increasing medical interventions which affects the immune system to treat numerous conditions in the aging population with increasing life expectancy.  

Risk of reactivation is stratified into high, moderate or low risk depending on the HBV status and type of immunosuppressive treatment (refer to reference 2 for details).Based on this risk assessment, reactivation of HBV can be prevented with eitherprophylaxis or pre-emptive therapy (through close regular monitoring) with effective antivirals.

Therefore, as antivirals are potentially prescribed in OBI for certain immunosuppressive treatment to prevent reactivation, it is important to establish if indeed the patient has had true past HBV exposure.

Most would rely on screen positive anti-HBcore in the absence of sAg as indicator of OBI prior to immunosuppressive treatment. Beware that isolated core positive result can also be due to a false positive in the test assay (specificity of current assay is ≥ 98%) or if the patient had received blood products which could contain anti-HBcore.

It is important to review and look at previous results for evidence of HBV exposure (previous serological markers) and establish history of prior HBV vaccination. This will guide test selection and help with result interpretation.

Follow the algorithm for screening below for those who had not been screened recently and not vaccinated:

Note:

*Specificity of test assay is ≥98%, therefore there is a 2% incidence of false positive results.

Reference:

1. Ang LW et al Epidemiology of hepatitis B virus infection in Singapore.. Epidemiological News Bulletin. 2010; 40 (4): 76-87

2. Loomba R, Liang TJ. Hepatitis B reactivation associated with immune suppressive and biological modifier therapies: current concepts, management strategies, and future directions..Gastroenterology 2017; 152: 1297-1309.

3. Gutiérrez-García ML et al Prevalence of occult hepatitis B virus infection.. World J Gastroenterol 2011; 17(2): 1538-1542.

We were recently sent a corneal specimen from a patient with suspected fungal keratitis.

On the KOH preparation of the specimen, broad hyphae with rare septa were seen (above and below).

Colony growth is typically white submerged colonies with few if any aerial hyphae (so it doesn’t look fluffy like a typical mould despite having hyphae).

5 days growth (front and reverse).

10 days growth (front and reverse). The radiate pattern of growth is most obvious in these images.

15 days growth (front and reverse).

Lactophenol cotton blue preparations of hyphae from colony growth.

Traditional culture-based identification of moulds is heavily dependent on observing the characteristic morphology of conidia (the asexual spores found on the tip of specialised hyphae).

So a lot of effort in the mycology lab is directed at getting moulds to sporulate and this used to account for a lot of the delay in lab diagnosis (you can imagine that it is not possible to identify a fungus by just looking at hyphae).

Because we were unable to get this isolate to sporulate, sequencing of the Internal Transcribed Spacer (ITS) was performed. This most closely matched that of Pythium insidiosum.

This is quite an important diagnosis to make.

Before the availability of ITS sequencing, the diagnosis of P. insidiousum was challenging and difficult, and it could also be under-reported because of lack of physician awareness (its still not in most microbiology tests). Knowledge of the clinical presentation with a high index of suspicion is required.

Though morphologically resembling fungi, P. insidiousum is not a true fungus and is more closely related to diatoms and algae. The cell wall is composed of a mix of cellulose-related compounds and glycan, and not chitin. Likewise, nuclei are diploid unlike fungi which are haploid.

This explains our difficulty with the morphological identification as formation of the characteristic oogonia (sexual stage) are rarely formed and zoospores requires special cultivation techniques using water agar and grass blades (Pythium spp. are usually plant pathogens).

Infections are associated with agricultural activity and are therefore rare in urbanised societies. However, they are well-described in the region particularly in Thailand and India (Pupabool J, 2006).

The most common infections are ocular as a result of direct inoculation. Vascular, cutaneous, subcutaneous and disseminated infections though very rare, have also been described. Vascular infections in particular are associated with thalassaemia and other haemaglobinopathies in Thailand.

Management is problematic as P. insidiousum  does not respond to the usual anti-fungal treatment. Treatment options that have been attempted include surgery, itraconazole and terinfine, potassium iodide, antibiotics like azithromycin and linezolid, and immunotherapy with P. insidiousum antigen (Agarwal S, 2018). 

Mendoza L, Hernandez F, Ajello L. Life cycle of the human and animal oomycete pathogen Pythium insidiosum. J Clin Microbiol. 1993 Nov;31(11):2967-73. Erratum in: J Clin Microbiol 1994 Jan;32(1):276. (free access)

Pupaibool J, Chindamporn A, Patrakul K, Suankratay C, Sindhuphak W, Kulwichit W. Human pythiosis. Emerg Infect Dis. 2006 Mar;12(3):517-8. (free access)