SIBO SERIES PART 2: The Science (and Art) of SIBO Testing

We know that SIBO is hugely underdiagnosed and potentially responsible for up to 85% of IBS cases (1); people who are told that there is nothing identifiably wrong with them. But how can this be? Why is SIBO so commonly missed or overlooked? A big part of the problem has to do with SIBO testing; who to test, when to test, how to test and most importantly how to interpret the SIBO breath test results. This is an area of SIBO research that has changed quite a bit in the last few years and one that many practitioners haven’t kept pace with. For a lot of IBS sufferers, a proper diagnosis is so critical, both psychologically and for informing effective treatment to overcome the horrible symptoms that negatively impact on your overall health… something that can only be achieved with the right testing.

Back in 2014 someone very close to me was suffering from hideous diarrhea, bloating, nausea, weight loss and stomach cramps. She was so clearly ticking all the boxes for SIBO so I asked her to request a breath test via her family GP. He hadn’t heard of the test but was eager to learn and so the test was completed with an interpretation from the lab; a massively incorrect interpretation, using out-of-date reference ranges, that had this dear person returning to me feeling defeated because she didn’t have SIBO. Now, I’m not here to play the blame game, I just like to illustrate that the science is ever-changing and after a quick glance over her results it was obvious, she did in fact have a raging overgrowth in her small intestine. The other clue - her symptoms; which improved dramatically once she started treatment. I’m hoping that last week’s blog (Part 1) has you clued up about exactly what SIBO is and maybe even some insight about what caused it for you. This week let’s jump into SIBO testing!

Here is a summary of what we are going to cover: Part 2:
> The ‘gold-standard’ SIBO test you need to ask your practitioner about
>  Why a negative test result doesn’t always mean you don’t have SIBO
>  Why SIBO testing and SIBO retesting is so important
>  Other GI testing you should consider

Refresh: SIBO SERIES PART 1: Is SIBO The Hidden Cause of Your IBS?
Jump to: SIBO SERIES PART 3: Latest SIBO Treatment Protocols That Actually Work


The ‘gold standard’ for non-invasive SIBO testing is a breath test that measures the levels of hydrogen and methane gas in your breath (2). I use the SIBO Lactulose and Glucose test by SIBO Test with my Australian clients and most commonly BioHealth Laboratories for US clients. It is a simple, non-invasive test that can be ordered online and completed at home.

After a 1-2 day prep diet, a baseline breath sample is taken before a lactulose solution is swallowed and breath samples are collected every 15-20 minutes for 2.5 to 3 hours (depending on the lab used), so 10 samples in total. The following day, the same process is completed with the glucose solution. If bacteria are present, they will ferment the lactulose or glucose and produce hydrogen and/or methane gas, which we can then measure through your breath.

In simple terms, if the gas levels are above a certain number within a certain time period, then the test indicates a positive result for SIBO. Using the correct solution and interpretation of results is a science all and of itself. Having a practitioner, skilled in the interpretation of SIBO breath tests is key to getting the right diagnosis. See below for a sample of what test results look like.

SIBO (Lactulose Only) Breath Test Result - Example Extract

SIBO Breath Test Result Methane Hydrogen Lactulose

SIBO Breath Test Result Methane Hydrogen Lactulose


As we’ve already touched on, the interpretation isn’t always black and white. With that in mind, here are a few different measurements (but not all) we are looking for that may indicate the presence of SIBO:

Lactulose Test:
> A rise of 20ppm in hydrogen above baseline within 90-120 minutes - Hydrogen Dominant SIBO.
> A rise of 12ppm in methane above baseline within 90-120 minutes - Methane Dominant SIBO.
> A combined rise in hydrogen and methane of 15ppm above baseline within 90-120 minutes - Mixed SIBO.
> Elevated baseline methane level that remains high (above 12ppm) - Methane Dominant LIBO (Large Intestine Bacterial Overgrowth) or potential H.pylori infection.
> Flat-lining (below 3-4) of both hydrogen and methane gas for the entire duration of the test period - Suggestive of Hydrogen Sulfide SIBO.

Glucose Breath Test:
> A rise of 12ppm in hydrogen or methane within 90-120 minutes - Hydrogen/Methane/Mixed SIBO.


The lactulose breath test is generally favoured within the SIBO practitioner community and is the substrate solution I most commonly use with clients. The reason we use lactulose is that it is scientifically proven to travel through the full length of the small intestine to the distal portion where SIBO typically likes to hide; i.e. close to the large intestine (3). There is limited research at this time about whether glucose has the same ability to reach the distal part of the small intestine.

However, numerous studies have shown the ability for lactulose breath tests to be interpreted incorrectly and clients given false positive results (4). That is, they are told they have SIBO when they really don’t. How does this happen? It’s all do with transit time; how fast the lactulose moves through your small intestine and into your large intestine.

A quick refresh from Part 1; while a huge amount of bacteria live in the large intestine, SIBO occurs when bacteria move to where they don’t belong and colonise the small intestine, a region that should contain very limited numbers of bacteria (5). So ideally, gas levels in the small intestine will be low, while they will be higher in the large intestine where all the bacteria live and feed on the lactulose solution. The breath test assumes that the lactulose solution that you drink will not make it’s way through to the large intestine in the first 90 minutes of the testing period.

But, for those with an anatomically shorter intestine or faster transit time (often those with diarrhoea-dominant symptoms), if the lactulose makes its way through the small intestine faster than expected, then you’ll likely get a false elevation of gas. When looked at in isolation and without a full history of symptoms and other variables, a practitioner might diagnose you with SIBO, when in fact, the elevated gas levels are coming from the large intestine, rather than the small intestine.

This is where the art of interpreting SIBO breath testing comes into it. It’s not always black and white. If you experience diarrhoea-dominant symptoms and only register an elevated and sustained hydrogen or methane result at the back-end of the testing period, then this may increase your risk of a false positive result.

This is one of the reasons why there is a lot of fear surrounding SIBO treatment. Lots of people have heard or read that you can never get rid of it; of people repeatedly treating it, retesting and still showing (false?) positive test results. And while for a small subset of people, just gaining symptomatic relief can be a long process, for the majority of clients SIBO is something that can be overcome. But if you don’t understand how to 1) treat SIBO and 2) interpret test results, you can fall into the trap of believing you’ll never be able to get rid of it!

The concept of false positives is really important for you to understand so that you don’t feel like there is something wrong with you or that you have failed somehow in taking the antimicrobials or eating something you shouldn’t have. It also means that if you’ve improved symptomatically, then there might be no need to continuously be treating something that doesn’t actually exist, but is an error with interpreting your test results. 


Glucose is often favoured in the scientific literature because of the requirements for strict (black and white) interpretation guidelines. While lactulose is prone to false positive interpretations, glucose is more prone to false negatives (6). That is, you are told you don’t have SIBO, when really you do.

This happens because, unlike lactulose, glucose is rapidly absorbed in the proximal (initial part) of the small intestine. If your overgrowth has made its way up to this part, then the glucose test will pick it up and yes, you definitely have SIBO. But, given that most SIBO overgrowths occur closer to the large intestine (in the distal part of the small intestine), the glucose might never make it down this far and the test may miss a large number of SIBO sufferers as a result. That said, clinically some practitioners are noticing that glucose is in fact producing positive test results after 90-120 minutes, indicating the solution has likely made it to the distal part of the small intestine or even into the large intestine.

For this reason, and because different bacterial species feed on glucose versus lactulose, the best way to be sure is to test using both solutions and to rely on a practitioner skilled in the nuanced science of interpretation.


Another reason you might get a false negative is if you have the other, less common or understood type of SIBO; hydrogen sulfide SIBO. There is currently no breath test available for detecting hydrogen sulfide SIBO which makes diagnosis difficult. However, flat lining of methane and hydrogen on a lactulose breath test (below 3-4 for the duration of the test), when symptoms of smelly rotten-egg gas are present, can indicate the presence of hydrogen sulfide SIBO and should be treated accordingly.


While breath SIBO testing is by far the most commonly used in clinical practice, there are a number of other other indirect ways to screen for SIBO. The first is a relatively new method that involves testing for anti-vinculin antibodies (discussed in Part 1) (7). While not technically SIBO-specific, a positive test result indicates that a client has had a significant exposure to a bacterial toxin that has most likely affected their migrating motor complex. A negative result means that you may still have SIBO, just not from an autoimmune trigger.

Another common way to screen for the risk of SIBO is through a GI Map or Comprehensive Digestive Stool Analysis (CDSA) test. There are a number of markers to look for, including calprotectin, elastase and fat in the stool that indicate SIBO may be present. While a CDSA test is not a cost-effective or reliable way to definitively test for SIBO on its own, given that most clients are also testing for parasites, bad bacteria, candida, h.pylori as well as other pathogens, gut function and levels of good bacteria, it is a sensible way to also determine whether a SIBO-specific test is warranted as a next step.


The simple answer is “test, don’t guess”. Diagnosing SIBO through anything other than a breath testis not recommended and prone to error. Not only in determining whether you have SIBO, but also how bad and what type - hydrogen or methane.

In a lot of cases, overgrowths are quite severe and multiple cycles of treatment are required to eliminate the bacteria entirely. SIBO testing gives us a great baseline measure to see how much treatment you might need. Similarly, retesting after each round of treatment is critical to understanding progress and to ensure the bacteria are fully eradicated. Testing also helps psychologically; just knowing you are heading in the right direction is a massive motivator.

Whether your results show elevations in hydrogen, methane or both are also important for determining the type and duration of antimicrobials to take. And for those using a phased approach to SIBO treatment, retesting is key to understanding when to make changes in diet and when to introduce prokinetics and other supplements into your protocol.


So, that’s pretty much everything you need to know about SIBO testing. Here’s a summary of things to remember:
> The gold-standard of non-invasive SIBO testing is a 2.5 to 3-hour hydrogen and methane lactulose and glucose breath test
> Correct interpretation of SIBO breath tests is important to avoid false positives and false negatives
> SIBO testing and retesting are really important to understanding not only whether you have SIBO, but also how bad and what type.

CLICK HERE to learn more about working with our team if you are interested in getting SIBO testing completed and/or interpreted.

Refresh: SIBO SERIES PART 1: Is SIBO The Hidden Cause of Your IBS?

Jump to: SIBO SERIES PART 3: Latest SIBO Treatment Protocols That Actually Work


  1. Bures, J., et al. (2010). Small intestinal bacterial overgrowth syndrome. World Journal of Gastroenterology : WJG, 16(24), 2978–2990

  2. Saad, R., Chey, W. (2014). Breath testing for small intestinal bacterial overgrowth: maximizing test accuracy. Clin Gastroenterol Hepatol, 12(12), 1964-72

  3. Posserud, I., Stotzer, P., Björnsson, E. S., Abrahamsson, H., & Simrén, M. (2007). Small intestinal bacterial overgrowth in patients with irritable bowel syndrome. Gut, 56(6), 802–808

  4. Simrén, M., & Stotzer, P. (2006). Use and abuse of hydrogen breath tests. Gut, 55(3), 297–303

  5. Gorbach SL. Microbiology of the Gastrointestinal Tract. In: Baron S, editor. Medical Microbiology. 4th edition. Galveston (TX): University of Texas Medical Branch at Galveston; 1996. Chapter 95

  6. Ghoshal, U. C. (2011). How to Interpret Hydrogen Breath Tests. Journal of Neurogastroenterology and Motility, 17(3), 312–317

  7. Pimentel, M., et al. (2015). Development and Validation of a Biomarker for Diarrhea-Predominant Irritable Bowel Syndrome in Human Subjects. PLoS ONE, 10(5), e0126438.