• We cover multiomic identification: unveiling the identities of bacteria, yeast, and fungi from all angles.
  • Genotypic Analysis: Deciphering genetic codes with Sanger DNA Sequencing. Sequencing of ribosomal RNA regions of bacteria and fungi.
  • Proteotypic Profiling: Exploring protein landscapes through Bruker MALDI-TOF. Analysis of ribosomal proteins.
  • Phenotypic Insights: Odin’s lens into microbial behaviors. Analysis of biochemical reactions, acids and salt tolerance, metabolism, fermentation, etc.

Learn more during our webinar, read the transcript below, or reach out to speak to someone today.

Transcript:

Hello and thank you for being with us today. My name is Brian. I’m currently the lab manager at Biolog Lab Services. Today we will be talking about multiomic microbial identification, utilizing our in-House services.

Multiomics is proving the old saying, “the more we learn, the more we realize how much we don’t know.” While there are different omics methods, using the single-omic approach has been the way to go for most researchers. However, the single approach limits the insights into microbial behavior and characteristics, only capturing a small subset of the biological cascade.

Instead of viewing biology through a single lens, the multiomic approach allows researchers to be able to obtain a more rounded view of microbial species, capturing the compass interaction between genes, their expression, and the proteins.

At Biolog, we offer three different omic approaches. 

  1. Genomic with our 3,500 XL DNA Sequencer
  2. Proteomic with our Bruker MALDI-TOF process 
  3. Phenomic with our own Biolog machine, Odin

We will be discussing these three different methods during the presentation and showing you a case study performed via all three methods. 

We do genomic testing by using Sanger Sequencing. What are some of the advantages? Some of the advantages of standard sequencing are high accuracy in determining DNA sequences. It can also sequence long fragments of DNA to enable the identification of specific genomic regions. It has been a cornerstone of microbial genomics research for many years and is referred to as the golden standard.

Here at Biolog Lab Services, we offer the genomic approach. With the genomic approach, we can identify bacteria using the 16S region at 500 bp and 1500 bp. For yeast and fungi, at 300 bp. All the way up to the species level. We do offer different turnaround times, from same day sequencing all the way to our three-day standard turnaround time, to accommodate all our customers.

Here’s an example of our DNA sequencing report. You will receive the top 10 matches with the genetic % difference and the length of the DNA. Our data analyst will make a call to the closest match and give it a confidence level based on the genetic % difference. The neighbor joining tree displays the interspecies relationship between the top 10 matches and the unknown. The distance between the matches is the genetic % difference.

We do proteomic testing by using Bruker’s ‘matrix assisted laser disruption ionization time of flight’, also known as MALDI-TOF. What are some of the advantages? Some of the advantages of MALDI-TOF is how rapid and reliable it is. It can accurately determine the identity of a microorganism from culture in a matter of minutes, rather than the hours or days needed for other methods. It uses a mass spectrum obtained from an organism’s protein fingerprint and compares it to a database to identify the organism, allowing researchers to achieve structural identification and characterization of proteins and peptides. It is compatible with many sample types, allowing for flexibility for experiment design.

Here at Biolog, with the proteomic approach using MALDI, we can identify bacteria, yeast, and fungi up to the species level. We also offer different turnaround times from next day MALDI, all the way to our three-day standard turnaround time, to accommodate customer needs.

Here’s an example of a MALDI report. You will receive the top 10 matches with a score value that is calculated by a matching algorithm, comparing protein spectral patterns. Based on the score value, we can assign a confidence level to the top match. The score value key is also provided on the report to allow for easier interpretation.

Now I will let Max, one of our Field Application Scientists at Biolog, talk to you about our Phenomic approach using Biolog’s Odin system.

Thanks, Brian. Our phenotypic ID system works by comprehensive phenotypic profiling. Odin enables you to simultaneously assess many different phenotypic traits, including metabolic activities, different growth kinetics, as well as environmental responses. We use this information to detect, or accurately identify your organism, based on the subtle differences between phenotypic traits among the microbial strains. 

With Odin, for microbial identification, we can cover a wide range of different types of species, including aerobic/anaerobic yeast and filamentous fungi. In this table, we can see a few different examples of the types of organisms, as well as the time it took to get to a confident identification.

For our bacteria up there at the top of the table, most of those were able to be ID’d within five to nine hours. S maltophilia took the longest, with a range up to 13.5 hours. Next with the YT or yeast plates, our Candida species took 48 hours to ID, whereas K marxianus was 72 hours. Our filamentous fungi were able to identify in as little as 24 hours, depending on the species, but some can take up to 96 hours.

And then finally, with the anaerobe plates, these all identify within the same amount of time, because we incubate them offline in an anaerobic environment. We then read them once on either the MicroStation or the Odin instrument, to be identified within 22 hours.

So, how does this all work? We have specialized plates for fast microbial identification. Each well will contain a different substrate, either a metabolic substrate or an inhibitor, as well as the redox dye that responds to energy production. What this does is create a color development in a pattern of wells that we can use as a phenotypic fingerprint in order to identify species or even strains from one another.

Our ID database covers over 1500 aerobic taxa, 360 plus anaerobes, more than 267 yeast species, and 700+ filamentous fungi. We also offer you the ability to develop your own local custom database, just in case your organism of interest is not in our database. We don’t hang you out to dry there. You’re able to identify it in other ways and also, you’re able to identify down to the strain level, as I mentioned. These are unique taxa that are included in our database, which includes different clinical isolates of some species, or environmental isolates of others.

This is all able to happen because of our superior chemistry where we incorporate more tests than different clinical phenotypic ID systems, which only use 20 to 48 tests, which allows us to have higher resolving power and more confidence in our identifications.

And finally, we have this very simple test procedure that takes only about a minute and requires no pretest, no gram stains, no colony morphology, no measuring, and no counting cells. You just pick a colony, put your swab into the solution, and then inoculate a hundred microliters per well; and that’s it.

And what is a study without evidence? As mentioned before, we did a case study of a known sample, Staphylococcus epidermidis, using the multiomic approach with all three techniques mentioned during the presentation. This is a sample of what it’s like to run an organism through all three services, giving you a well-rounded view of the organism.

With Sanger Sequencing, we’re able to see the genome sequence and we’re able to provide an effective file if requested. With MALDI, we’re able to see the activity of proteins and peptides. We’re able to provide a mass spectrum if requested. With Odin, we can access multiple phenotypic traits, including metabolic activities, grow kinetics, and environmental responses.

After running our Staphylococcus epi sample (09:17) through all three methods, we obtained our reports. For MALDI-TOF. We got the top 10 matches. As you can see, all top 10 matches came out as the same genus and species, with the top three matches giving us a species level. The sources shown are the different culture collections where the data was obtained, even though we do not have the capabilities to ID to the strain level, all these sources show that our sample is a Staphylococcus epi.

The score value is different for each, but this could be due to the cultural environment where these strains were tested at. The top source seems to be like the test environment, where we perform MALDI. With Sanger Sequencing, we also get our top 10 matches, with the top 10 matches having a 0% genetic difference from the known organisms. Since the second top match has a % genetic difference, less than 1.2% and it has a different species, it gives us a species closely related confidence level.

Why does this happen? Since DNA is the backbone of the organism, all these different species are genetically related. Therefore, through 16S DNA Sequencing, we are not able to differentiate the species. However, if we go back to MALDI, we can see that through proteins we can confirm the species of the organism.

The neighbor joining tree shows us the interaction and distance between the top 10 matches. As you can see, Staphylococcus epi has the shortest distance to our sample.

Next, for phenotypic ID, when we tested Staph Epidermidis, it was again our top result with a similarity of 0.5, which meets our cutoff. You can see below that the next ranked microbes, which are a closely related species. This emphasizes the fact that phenotypic ID can oftentimes give you a unique profile based on your specific species or strain. On the right-hand side, you can see the expected profile of the strain with the false positives or negatives, depending on what the algorithm was expecting.

As mentioned before, you’re able to gain a lot of functional insights from this phenotypic ID, as well as the identification. Now that we’ve confirmed the identity of the Staph Epidermidis through three different methods, we can now learn a little bit about the organism. Based on our Gen III plate results, we now know that S. epidermidis prefers certain carbon sources over others, including dextrin, maltose, fructose, glucose, pectin, methyl pyruvate, lactose, etc. It’s also resistant to some stressors that other bacteria may not be, including: pH6, Na Cl, sodium lactate, tetrazolium violet, LiCl, nalidixic acid, Aztreonam, sodium butyrate, potassium tellurite, etc.

Here’s a view of all three final reports. Even though each technique approaches the organism from different angles, in the end, they all agree on the same result. This confirms that our sample is in fact S. epidermidis. Integrating these three different methods allows for a more well-rounded view of the microbial species. This captures the complex interactions between genes, their expression, and the proteins. This gives us a more accurate microbial identification. And when we have a holistic understanding of microbial organisms, this allows for the development of targeted interventions, such as antimicrobial treatments that can be tailored to specific microbial characteristics.

Here at Biolog, we’re a multiomic microbial identification testing lab, striving to pour our focus into our customer’s needs. We have a plethora of different services within a scope of genomics, proteotypic, and phenotypic analysis. This allows us to provide a complete polyphasic approach to our research and services. You can always contact us if you’re interested or have any questions about our services. 

Whether you are concerned about contamination, enumeration, polyphasic, genotypic, proteotypic, or phenotypic test; Our testing services can help you reduce costs and guarantee brand safety. Biolog Lab Services results are ISO 17025: 2017 accredited and validated. We are registered with the FDA, GDUFA, and cGMP compliant. These accreditations prove that our lab has an acceptable quality management system in place, plus the ability and competence to provide testing and calibration results.

Here are some of the most common samples we receive in our lab, but we can receive lots of different types of samples depending on your needs and workflow. Feel free to contact us if you have any questions about what type of samples you can send, as well as how to ship them to us. This applies to both domestic and international customers.

As we come to the end of our presentation, let me introduce you to our ecosystem. At Biolog, we are on a mission to shed light on the intricate world of microbes and cells. Through cutting edge technologies and pioneering analysis, we offer fit for purpose instruments, a comprehensive database, custom plates, and accredited multiomic lab services. Think of us as your partner, propelling your scientific pursuits forward.

Thank you for joining us on this enlightened exploration.