Speaker: Bryan Turner:

Hello, everyone, and thank you for attending today's webinar: Rapid Molecular Diagnostics for Sexual and Women's Health. I'm Bryan Turner, a marketing manager for Cepheid, and I'll be introducing our speaker, Amery Vilander. Amery serves as the microbiology lab manager for UC Health Memorial Hospital in Colorado Springs. She participates on several teams within UC Health, including the microbiology molecular best practices team, the South Region Antimicrobial Stewardship Work Group, and the Infection Prevention Work Group. Amery has extensive experience with all facets of new molecular test implementation in a hospital laboratory, from conducting the verification process, insuring CAP compliance, to keeping microbiology lab staff engaged. In 2021, Amery earned the Clinical Excellence Award for transformation leadership from UC Health Memorial Central. She welcomes the opportunity to share with others her experiences in onboarding new tests and best practices she has implemented at UC Health, and now I will hand it over to our speaker.

Speaker: Amery Vilander:

Okay, so the objectives for today, first we're going to discuss the most recent CDC surveillance data for sexually transmitted infections. I'll also explain the benefits of rapid, on-demand molecular diagnostics compared to traditional methods like culturing, and wet preps, and of course batch testing. We'll become familiar with common molecular testing options for group B strep, chlamydia trachomatis, neisseria gonorrhoeae, and vaginitis. And then lastly, we'll talk about some key considerations that you'll want to think about when you're implementing a new molecular diagnostic assay in your laboratory. Today's disclosures. I do have an honorarium for this presentation provided by the Cepheid Speakers Bureau, and a couple of abbreviations there. Really long names for organisms, so I'm going to use CT for chlamydia trachomatis and NG for neisseria gonorrhoeae. I may also call it GC as well, for gonococcus.

The CDC surveillance data, in 2021, this was preliminary data. CDC says 2.5 million reported cases of chlamydia, gonorrhea, and syphilis. Now, if you do the sexually transmitted disease testing in your laboratory, you may have noticed over the last several years, an increase in positivity for your tests. It is something that we noticed over the last few years, and CDC's last report was published in 2020. It is something that they report on every year, and in 2021, they have released some preliminary data. Something to consider for surveillance data is that this report for 2021 is going to be finalized probably later this year, and the '21 report ran data through the fall of 2022.

So the impact of COVID on sexually transmitted infections, you may be wondering what happened during this time. Even though we've been seeing kind of the steady march of increase in infections over the last few years, we would theorize that maybe something declined a little bit during COVID. In actuality, case counts in 2020 were all higher than 2019. Now, they did fall in the early days of the pandemic, in March and April. So you'll see here that, for gonorrhea and syphilis, it was a very steady, and sometimes a very steep increase, in case detection, but you'll notice, for chlamydia, kind of that sharp drop there, and what CDC is saying to explain that drop in chlamydia was kind of multifactorial.

There were reduced preventative screenings during that time. The shift in resources to COVID, and really looking at symptomatic patients, and something about chlamydia is that oftentimes, patients are asymptomatic, so the detection of the positive cases is really done through a routine preventative screening. Obviously, social distancing was also limiting sexual activity, so overall, with these reasons, CDC says that that is why chlamydia was disproportionately affected, compared to GC and syphilis. And chlamydia actually had a 13% drop by the end of 2020, so again with that sharp decline. But you can tell by 2021, things were starting to rise as the communities started to reopen.

For drug resistant gonorrhea, or GC, there is a CDC report. It's called the AR Threats Report. It was last published in 2019, and there are actually three categories, organisms, for this report, primarily bacteria and yeast. The first category is concerning, second category is serious, and then the top category is urgent. And you'll notice that the CDC has actually put drug-resistant neisseria gonorrhoeae, or GC, into that top category for concern. 550,000 drug-resistant infections every year, 1.14 million total new infections each year, and several hundred million in terms of lifetime direct medical costs. On the right here, we have this graph that shows emerging antibiotic resistance over the last 16 years, and kind of the timeline for how treatment has evolved over time. Currently, ceftriaxone is the recommended drug of treatment.

For pharyngeal infections, the CDC does have STI treatment guidelines. They were updated in 2021. The last time they were updated was 2015. And something about pharyngeal gonorrhea is that it's largely asymptomatic, and it is common in certain patient populations. It is harder to eradicate compared to urogenital or rectal, so this is something that maybe not a lot of people know about, when you have gonorrhea in the pharyngeal area, that you actually want to have a tested cure seven to 14 days after initial treatment, and again, three months anywhere up to 12 months afterward.

NAT, nucleic acid amplification testing, is superior to culture in that regard, and then CDC says, in these STI treatment guidelines, that you do not need to retest if your infection is in the urogenital or rectal area, but pharyngeal, you need to treat and then test for cure, because it's hard to eradicate. So when you have a suspected treatment failure, you should have both NAT testing and culture, and then if you grow it upon culture, you should do susceptibility testing. And I realize not a lot of labs out there will do automatic AST testing on a neisseria gonorrhoeae isolate, so that's something you'll want to send to your reference lab if there is a request for that.

Now, with pharyngeal infections of gonorrhea, not all assays have that as a specimen type in their assay, so you will want to read the instructions for use, or the package insert more commonly called. Sensitivity and specificity are going to vary for pharyngeal, potentially, versus urogenital or rectal. And then some assays do have cross-reactivity with your commensal or like your normal flora neisseria, species that are in the pharynx. These are things to consider when we are talking about pharyngeal infections for gonorrhea.

The CDC surveillance data for group B strep and vaginitis. I'll talk about group B strep first. The CDC does have an Emerging Infections Program, and this is a program that encompasses multiple states across the country. You may be in a state that participates in this. I know Colorado is one of them. In 2020, they reported on the national estimates of invasive disease, and what we define as invasive disease is when group B strep is found in a sterile body site, such as blood or spinal fluid.

So you can see early-onset cases there, late-onset cases, total cases, and deaths. This is a pathogen that is very capable of causing serious harm, and the references there at the bottom, it's a wonderful report if you would like to take a look at it. It actually atomizes by state if you'd like to take a look at that. This is why it's important and in CDC guidelines to test for group B strep.

For vaginitis, we're really talking about bacterial vaginosis, candidiasis, and trichomoniasis. The CDC has national estimates of disease, and the statistics are pulled right off the CDC website, with the references listed below for you there. Bacterial vaginosis is the most common vaginal condition in women between the ages of 14 to 49, 29.2%. Candidiasis is the second most common vaginal condition, with an estimated 1.4 million outpatient visits every year. And trichomoniasis, 2.6 million women, 2.1%, ages 14 to 49. For trichomoniasis, it is caused by a parasite, and it is largely asymptomatic, 70% of the time. Women are more likely to have symptoms than men.

All right, next we will talk about rapid diagnostics versus batch testing and traditional methods. The considerations for, if you want to bring up a rapid molecular test versus something else or maybe sticking with your method that might be more traditional, like culturing or wet prep. It's not always an easy decision. Sometimes, it's something that you're asked to do from maybe your medical director, or maybe your customers there in your hospital, or your community. I kind of put together a slide here, of things to think about, right? If you were going to think about doing rapid molecular diagnostics, obviously the benefits are turnaround time. It is going to be faster than waiting for an organism to grow. You are going to have better sensitivity, meaning you're going to pick up, potentially, small numbers of organisms that maybe you don't even get to grow on culture.

And then, consider that molecular diagnostics, especially in the setting of CT and NG in emergency departments, if they have the result, it helps to avoid over- or under-treatment. So there's a great study there I've listed at the bottom, of a randomized clinical trial, where they looked at CT/NG in an emergency department for women, and what they realized was that if they had patients that had the rapid diagnostic done, they have the right treatment, and correct meaning correct to that result, right? If they didn't have it, they would either be under-treated or over-treated, so I think that's a really great study to look at if you're kind of trying to figure out, "Do we implement CT/NG in the emergency department, for emergency department patients? Do we batch it? Maybe we run it 24/7."

The other things to consider for rapid molecular diagnostics are what are your test volumes expected to be, and your instrumentation, and your staffing. Obviously, that's important to make sure that you have the infrastructure to be able to accommodate a rapid test volume, especially if it's significant. You'll want to think about what kind of resources you have for doing a verification study, and implementing that, and really managing that project. There are supply and labor costs to consider, and you'll want to get input from your stakeholders. These are the people who are involved, not only with running the tests, but also who order the tests. This might be looking at who is it that would potentially order this test? It could be your emergency department. It could be your women's clinics, or your OB/Gyn, possibly your primary care clinics and urgent cares, so those are things that you'll want to consider and get buy-in from them, and let them know that this might be an assay that could be of value.

And then if you're part of a system, you know, a larger health system, and you have some guidelines in place to standardize practices, or maybe you have an existing agreement with a current instrument platform, these are all going to roll into your decision of timing, and when we switch, or do we switch? And of course, test complexity and personnel requirements. A lot of molecular testing is moderate complexity, but some of it is high complexity, so you'll want to consider what kind of personnel you have and the requirements for your assay.

[inaudible 00:15:09] right? Let's talk about, maybe I decide not to do rapid molecular. Okay, so maybe I decide, "Do I want to stay in culturing? Do I want to stay on wet preps?" Like I mentioned in the setting of vaginitis. Maybe we want to batch our assays. Obviously, there are some benefits to doing that. Your supply costs are typically way less, and you may have some fewer regulatory requirements if we're talking about sticking with cultures and wet preps, in terms of you wouldn't need to do a verification study.

But some things to consider for a traditional method is that if you're going to go with culture, even it's in the setting of group B strep, or chlamydia, or GC, some of these organisms take several days to grow, organism identification does require high-complexity testing for personnel, and your cultures are going to incur additional charges for organism ID. And then in the setting of chlamydia, not only does it take a long time to grow, you need special media in order to do that. So largely, for CT/NG, and also for some of your vaginitis targets, this is where maybe molecular may come in and be a better choice for you. And for BV, bacterial vaginosis, Gardnerella vaginalis and lactobacillus are actually not specific, due to being found in healthy women. There is a study there by Fredricks, that talks about how Gardnerella vaginalis can be found in up to 70% of healthy women, and I will say that when we get to a later part of the presentation, it is something that we kind of noticed in our previous assay about, "Wow, why are we seeing so much Gardnerella?" That started the wheels turning in our head initially, but it started with that.

Group B strep, non-beta-hemolytic strep B, can be missed on culture, so I believe it's about 20% of group B strep can actually be non-hemolytic, so that means if you're screening for your cultures and you're looking just for beta-hemolysis, there is a possibility that you would miss group B strep on there, especially if there's a lot of vaginal flora growing on that plate. And of course, for trichomoniasis, on a wet prep, you need to visualize the movement, that kind of jerky movement, in order to be able to report the trichomoniasis. Typically, it's about a one-hour time before they start to die. So, considering, "Do we go rapid molecular? Do we stay with our current method?" These are things to consider.

All right, so rapid molecular diagnostics. We'll be talking about GBS, group B strep, CT/NG, and vaginitis. We'll start with the group B strep rapid platforms. These are kind of the most common platforms. There may be others out there on the market. You know, these are the ones that are the most common, the Hologic Panther Fusion, Luminex Aries, Cepheid GeneXpert, and the BD Max. They will vary from being a floor-standing model to bench top model, roughly equivalent sensitivity and specificity among the models. For Cepheid GeneXpert, they do have a way to do two different assays, so I will say for all four of these methods here on the screen, they are all done on enriched vaginal-rectal swabs, in Lim Broth typically, and that is the method of detection that the CDC and ASM recommend for group B strep. As a matter of fact, there are catch-up list questions related to that.

Now, for the Cepheid assay, it is the only one that does have an option to do a direct method off of your vaginal-rectal swab, and that is typically for women in intrapartum settings, so they're already in labor and delivery. And that is a different assay, it's called the GBS assay, than the enriched assays. The enriched assay is GBS, LB, XC. It's typically for prenatal settings, and done on an enriched Lim Broth.

So there's actually a really good study out of UC San Diego there, from Dr. David Pride. He compares several methods for group B strep detection, molecular, and this quote is from an article. And really, it talks about how group B strep should be detected by NAT testing as the gold standard. Less hands-on time, definitely. You can throughput a lot of samples, especially if you have a patient population that sends a lot of GBS testing to you, faster time to result, and greater sensitivity. You're not going to miss some of the most hard-to-find GBSs that might be buried in the plate with other vaginal flora.

Next, I'll talk about CT/NG rapid platforms. Three most common assays out there on the market, Hologic Panther, the BD Max, and the Cepheid GeneXpert. Now, there are a variety of specimen times. Genital and urine tend to be done on all three. Now, extragenital sites may not be done on all of them. You are going to want to look at your IFUs, your package inserts. Sensitivity and specificity is going to vary by the organism, so you may have a different one, different sensitivity and specificity for CT versus NG. And then, just wanted to call out that Hologic and Cepheid have a CT/NG assay. The BD Max assay actually is a combo, and includes trich, so it's CT, NG, and trichomoniasis. Now, for all three of these, they are FDA cleared for vaginal self-collection in a clinical setting, or clinician collection.

So you may be wondering why is there a thing about vaginal-self collection? As explained to me by our ED partners, this is something I've learned more recently. It's not something that we really think about in the laboratory, right? You think, "Oh, the clinician collects the specimen," but for some patients who may have a history of sexual trauma, or they just would prefer to not have a clinician do a pelvic exam on them, the option to do a self-collection really is just such a welcome relief for some patients. So the sensitivity and specificity numbers that are there, those are for female urines, and then endocervical swab, so you can see that all of them have about 90% sensitivity, specificity for both targets.

For vaginitis, we've got four platforms here listed, again, the most common ones. There are some that have just one assay, meaning you're going to detect multiple targets in the one assay, and then for the Hologic Panther, they do separate out their assays, so you'll have to have a separate assay for bacterial vaginosis, and then a separate assay for candida and trichomoniasis. You'll notice on here, for the assays that do have Gardnerella and lacto, they're there in red, as I've mentioned, Gardnerella and lacto can be found in a large percentage of healthy women.

I will mention that the BD Affirm is not cleared for patient self-collection. The other three are, so you know, when we're talking about patients who have a history of sexual trauma, and they are needing and wanting to have a self-collection method, that kind of puts the ED in a difficult position to... if you're running an assay that's not cleared for patient self-collection. And then of course, for a pelvic exam in the emergency department, it often requires two providers in the room, so usually a provider and a second person to assist, so there's a benefit to the ED also, in terms of having self-collection available.

So you'll see, in terms of sensitivity and specificity, again, it's going to vary by organism. And we're talking about bacterial vaginosis, we're talking about yeast, and we're talking about some assays having Gardnerella, or even, when we were talking about a shift into anaerobic environments, we're talking about atopobium, megasphaera, BVAB-2, and that is its name, the name of the organism is BVAB-2. For those of you thinking, "Well, if it's an anaerobe, I should be able to culture it, right?" And these are not organisms that we typically culture. The vaginal specimens, not one that we typically do an anaerobic culture on. Notice that the Cepheid GeneXpert, their MVP assay does not actually detect Gardnerella or lactobacillus, and sensitivity and specificity listed there, on this slide, sensitivity and specificity are over 90% for the Cepheid assay.

Now I'm going to talk about the implementation of the Cepheid MVP assay in our laboratory. As I mentioned, I'm in Colorado. We are in Colorado Springs, which is the Southern region of UC Health, and in the fall, we moved to the MVP assay from our old method, which was the BD Affirm, and the story with MVP kind of began for us in about the early part of 2022. We were having issues with our current assay, and when I found out Cepheid was working on a vaginal assay, and I said, "You know, I think I need to explore this, because I know the hands-on time is really less than what we're currently doing, and it's a method that we're familiar with," and I knew that I needed to pull in the stakeholders to have that discussion.

So Memorial Hospital Central, we are a part of UC Health. I am sitting in that building that you see right there on the lower-left. We are in Colorado Springs. We are a award-winning level one trauma center, with a comprehensive stroke center. My microbiology lab team is comprised of 19 medical laboratory scientists, covering day and evening shifts. We do culture and molecular testing for four hospitals in the [inaudible 00:27:05] UC Health, and we knew that we wanted to switch methods, and this actually began pre-COVID, that we started investigating this.

So why change methods? When we realized that Cepheid was bringing up a new vaginal panel assay, we took a hard look at it and compared it to our current method, and our current method at the time, we were unable to run stats. We had to batch specimens, and we did not have random access, meaning I have a sample, I put it on the analyzer. I have another sample. I put it on the analyzer, to get it running. We did not have the capability to do that. The Affirm required us to do batching. It was a very labor-intensive test. It had many steps, different reagents you had to incubate in certain temperatures, and if you've ever run the BD Affirm assay, you have to squeeze the reagent ampules pretty hard, and talking about motor skills and having to really just squeeze, when you run so many of these samples through in 24 hours, that can present a challenge with ergonomics.

And then also, with the BD Affirm, there was subjective color interpretation. We had to look for rim dots, so then you're going, "Is it blue? Is it light blue? Is it sort of blue?" And this was not a test that we could interface. It was a test that required manual result entry, and then as I mentioned before, it was not FDA cleared for patient self-collection, so that really presented a challenge for our ED partners. So those were our challenges.

So when we moved to the MVP, kind of high-level overview with our wins. We reduced our turnaround time by 21%, and we have a Cepheid GeneXpert Infinity, but even with the Cepheid GeneXpert, you don't need to batch it. It is random access. We were able to interface it to our LIS, which is Epic Beaker, and we were able to set up auto-verification. Now, this is not something you currently do in your microbiology lab. I understand like the concept of it can be scary. I will say that just peruse it, pursue it. Like, ask your LIS partners, ask your chemistry hematology partners. Lots involved in that, because that has really changed how quickly that we can chart results to clinicians for patients.

And obviously, MVP has patient self-collection in a clinical setting, and I can't tell you how eager my staff were to participate in this. They were so motivated. They were wanting to run verification samples. They wanted to bring up this assay. And of course, we pulled in our ED partners, and said, "This is the assay. These are the targets that are going to report. You know, is this something that we can potentially move to?" And it was very warmly embraced.

So I'm going to talk about test implementation, some of the key points that you want to consider. Number one is to identify your stakeholders, and I can't stress this enough. It is going to be for your current tests, you should be able to figure out a way, either running a report or looking at your previously tested patients. Who is ordering your tests? Where are these specimens coming from? And since we're talking about women's and sexual health, group B strep, CT/NG, vaginitis, generally speaking, it's going to be your urgent cares and emergency departments, women's clinics, OB/Gyn, even your primary care clinics, things of those types of patients. So those are the potential stakeholders that you will want to have conversations with if you are considering bringing them a rapid molecular assay.

Of course, with your lab team, you're going to want to involve your pre-analytical, so those are the folks who are collecting specimens and processing your specimens, then of course your analytical staff. And maybe this is an assay that you maybe run in more of a general lab setting, or maybe you are a smaller facility and you run everything, you run all the tests, including microbiology. And of course, your lab leadership. It's going to involve a verification study. You will have to involve your medical director and any other medical director personnel, such as medical directors of emergency department or women's service lines.

Number two, you're going to want to manage this project. This is a big project, and if you need guidance, there are resources out there for you, but number one, check in with your lab quality team for guidelines, possibly from your quality management plan. You may have a document that kind of outlines for you the things that you need to do to bring them a brand-new test, so you'll want to write a list of tasks that need to be done, or maybe you have a checklist from your quality department. That's wonderful. That is something that we have in our quality department. And figure out who is responsible for that test, and then an approximate timeline.

And then, think about how many of the tasks are going to be going on at the same time. This is where this project management kind of kicks into high gear. Enlist help, because you will not be able to do it all. You are going to need help. You are probably going to be doing many of the same things, many of these things, at the same time, so obviously there will be LIS build, you'll have to test, or maybe it's somebody else that does testing. You will have to create a charge code, so you're going to have to figure out what your costs are to run the assay. You'll need to provide education to your stakeholders and your lab teams. You'll have to do training in competency, and then collection device changes, that sometimes is really challenging.

If you're considering an assay that is going to potentially have a change in collection device, you'll need to consider who it is that orders your collection devices. It might be you and the laboratory, and you supply them, or it might be your nursing partners that will order and stock those supplies in their respective areas. If that's the case, you're going to want to get education out to them in terms of, "This is the supply item you'll need to order. This is what it looks like when it arrives."

Regulatory requirements. Obviously, there's procedures. There's IQCP if you choose to run your assay and not do QC daily rotation testing. Obviously, there's CAP and proficiency testing. Now, maybe whatever regulatory body accredits your laboratory, they will have requirements as well. And then communication is key. You cannot over-communicate, and it gets very important to keep your stakeholders updated, especially if you experience any delays, education, just really open up that channel for communication to stakeholders and your lab teams.

Number three, since we're talking about organisms, group B strep, CT/NG, vaginitis, really involve your infectious disease and pharmacy teams, especially if you're changing targets, such as for vaginitis. Maybe you're going from an assay that has Gardnerella and lacto in it, and now maybe you're considering an assay that doesn't have it. There may be implications for what is used for patient treatment, based on the targets that are now going to be reported, so make sure you pull in pharmacy and infectious disease. And then number four, be proactive, but have a backup plan. For us, when we were implementing MVP, we knew, because vaginal specimens can be kind of an invasive procedure for a patient, and I would not want to have to do a recollect unless we absolutely had to. So for us, we made sure that we had a backup laboratory that could perform the old test, should we get a patient specimen on the old collection device.

So again, if you're talking about new collection devices. You want to order a lot of them, you want to order them ahead of go live. It's not something that you want to think about a week before go live. Like, this needs to be in your implementation plan. And you can arrange for a swap-out of supplies in clinics or labs if you're doing a change in collection device. So again, it's being proactive. It is a lot of work, and maybe this is something that someone in your inventory or supply chain can help you out with, but the more time you invest in that, I think it's going to result in fewer recollects needed after go live.

Check your IFUs. If there are additional devices in the IFU approved, include them in your verification. And what I mean by that is for example, if the IFU says that you can run it off a liquid amines or an e-swab, or you can run it off of a regular culture and swab, if you know that there is potential for your lab to receive both of those types of collection devices, consider including both of them in your verification study. That way, your team is not locked into only receiving one type of device. And again, check your IFUs, and only include what is included in your IFU.

Now, what if a clinic sends an old collection swab? This is a potential scenario. As I mentioned with the vaginal specimen, I hate to ask for a recollect on that, so you'll want to identify a backup lab. And this might be a reference lab for you, if you're more of a smaller facility and you don't really have anyone else to send to, or maybe you're a part of a larger health system, and there is somebody maybe that is going to keep the old assay for a little while. That might be an avenue for you to explore. And involve your LIS, because your LIS team may need to do some reworking of your test in order to do that. And we know that when a patient has to be recollected for a specimen, sometimes it does happen and it is needed, we have an opportunity to affect patient experience and satisfaction scores.

And then lastly, what if you get a wrong test order? Let's say you decide to change tests completely, and now you have not only a new collection device, but now you have a brand new test order as well. Who's going to educate to that, and how are they going to educate? This is obviously going to be tied into your LIS, and the education may not be done by you, or maybe it will be done by you. If you have an externally facing website that lists test collection information and ordering information, you'll want to make sure that you update that, if you have newsletters or you have like an email blast. Whatever method of communication you have for notifying providers of the new test, the new collection device, notifying your nursing partners, and again, being proactive but having that backup plan, is going to pay off for you if you're changing a test, or especially involving a new collection device.

So in summary, today we talked about CDC surveillance data, 2.5 million cases of chlamydia, gonorrhea, and syphilis in 2021, and that's preliminary data from the CDC. We also talked about group B strep, national estimates of disease, as well as vaginitis, vaginitis encompassing bacterial vaginosis, VDC, which is vulvovaginal candidiasis, and trichomoniasis. We talked about rapid diagnostics, benefits to doing that, and things to consider, as opposed to doing batch testing or remaining on culture or wet preps for your vaginitis. We talked about the most common molecular platforms for group B strep, CT, NG, and vaginitis, and then we also talked about key points for test implementation.

It is a large project, a lot of things happening at the same time, and I actually say that, as a laboratorian, running verification samples or just running QC is actually the easiest part of bringing the test live. There's so many other things that maybe not everyone fully thinks about when they are bringing a test live, and it is even more involved if you are changing a collection device, or changing the name of your test. I have all my references listed here. The IFUs can be found at various websites. You can take a look at all of the data that we talked about, and even the studies that were referenced in the presentation. And happy to answer any questions.

Bryan Turner:

Hello everyone and thank you for attending today's webinar. Group B Strep: A leading cause of early-onset neonatal sepsis. I'm Bryan Turner, a marketing manager for Cepheid, and I'll be your moderator today. Before we begin, we want to cover a few housekeeping items. At the bottom of your audience console are multiple application widgets you can use. If you have any questions during the webcast, you can click on the QA widget at the bottom and submit your question. We'll try to answer these during the webcast, but if a fuller answer is needed, or we run out of time, we'll answer later by email. A copy of today's slides are available in the resource widget at the bottom of your screen. You can also expand your slides by clicking on the maximize icon on the top right of the slide area, or by dragging the bottom right corner of the slide area. If you have any technical difficulty, please click on the help widget. PACE Credits are available for attending today's session.

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For those of you just joining us, welcome to today's webinar Group B Strep: A Leading cause of early-onset neonatal sepsis. Today we are joined by two speakers. Dr. Silverman, who currently works at the David Geffen School of Medicine at UCLA, where he has a faculty appointment as a Professor of Clinical OB-GYN, and serves as the director of Maternal-Fetal Medicine fellowship program. In addition to his clinical and teaching responsibilities, he's also the director of the department's new Infectious Diseases in Pregnancy Program. In that capacity, he's working with both adult and pediatric ID divisions, as well as the department of microbiology to develop a multidisciplinary program, to develop educational research in clinical care focus areas. Dr. Silverman has authored over 50 peer-reviewed manuscripts, 20 textbook chapters. And he has lectured widely with particular interest in obstetrics, infectious diseases, recurrent pregnancy loss, and prenatal diagnosis.

Our second speaker is Dr. Pride, who's originally from Nashville, Tennessee. He received his undergraduate degree in biology from Wake Forest University, his PhD in microbiology and immunology for Vanderbilt, and his MD at New York University. He is board certified in internal medicine, and received a subspecialty training in infectious diseases at Stanford. He's a professor in the Departments of Pathology and Medicine at UC, San Diego. Dr. Pride also serves as the director of microbiology laboratory at UC, San Diego Health. His major interests are in developing diagnostic tests for infectious diseases, understanding the role of microbial communities in human health and disease. His research laboratory specializes in human microbiome research for the focus on viruses that inhabit the human microbiome. His laboratory also focuses on understanding how to use bacteriophages to promote human health. And now with that introduction, I'm going to hand it over to our first speaker, Dr. Silverman. Take it away, Dr. Silverman.

Neil S. Silverman MD:

Thank you very much Brian. And thanks to all of you joining me from Los Angeles today to talk to you folks about Group B strep diagnosis and treatment. And to discuss the new guidelines that were issued about two years ago from the American College of Obstetrics and Gynecology regarding preventing Group B strep disease in newborns. My disclosure information is listed as you see it there.

So as background, what we're primarily going to be talking about is the guidelines that are designed to prevent early-onset GBS illness in newborns. Which is still the leading infectious cause of neonatal sepsis in the United States. This was first identified actually in the 1970s as a leading cause of sepsis. And in those early studies, maternal colonization with GBS was identified as the primary risk factor for the development of significant newborn illness. Before any of the guidelines were established back in the 1990s, the annual incidence of this illness was about two cases per 1000 live births in the United States. And since the guidelines have been introduced and refined over the past 30 years, that rate has been decreased to under 0.25 cases per 1000 live births in the United States. What we know is that the primary risk factor for neonatal early-onset disease related to GBS is maternal vaginal-rectal GBS colonization during labor.

Now, that's not to say that GBS in itself is a pathogenic organism in ordinary circumstances. We know it's a physiologic component of both the gastrointestinal and vaginal microbiome. And that the GI tract is the reservoir, the source, for maternal genitourinary colonization. However, colonization in this particular region in the body in general can be transient or persistent. And that studies have demonstrated that the prevalence of being colonized in pregnancy at any given time, including in labor, is about 30%. What we also know is that of the pregnant individuals who are colonized in the first trimester, only about a third of them will be still colonized at the time of delivery. So that's why the very early guidelines tried to develop a way to test folks during pregnancy in an appropriate manner that would be most predictive of their colonization status at the time of delivery.

Neonatal GBS illness is typically presenting on the day of life, or the first day of life. And 95% of these cases in infants occur within the first 48 hours. Early-onset disease in general is defined as illness that occurs within the first seven days of life. After seven days of life, it's termed late-onset disease. And that's actually a very different illness, and not what these guidelines are designed to address. Kids tend to present with sepsis, pneumonia, or less commonly, meningitis. Interestingly, the majority of these cases, almost three quarters of them, occur in full-term newborns. And GBS accounts for almost half of confirmed sepsis in this group of full-term babies. In the very early stages of identification of GBS as a source of illness, the case fatality rate, meaning the number of infected newborns who actually died, approached 50%.

Currently, with the introduction and adoption of management guidelines, about 2% of term infants will still have significant illness, including mortality, if they are colonized. But almost 20% of preterm infants will have mortality related to newborn illness. So it is an important thing to identify a risk for in all newborns, but particularly in preterm newborns who are more susceptible and vulnerable. This little diagram basically outlines the overall risk of mother to infant transmission of GBS. So if we start with a GBS colonized mother, about half of those newborns will be colonized and the other half will not. And of those who are colonized, the vast majority of them in a term infant will be asymptomatic and need no additional intervention. But 2% of them will develop early-onset sepsis, or pneumonia, or meningitis. So the maternal risk factors for early-onset GBS disease in newborns, we've already mentioned that the primary risk factor is maternal vaginal-rectal colonization. But also preterm delivery and extended duration of ruptured membranes, or water being broken of at least 18 hours. Or any evidence of clinical intra-amniotic infection during labor.

We also know that women who have asymptomatic bacterial colonization of their urine identified during pregnancy, are at much higher risk of maintaining that colonization as they approach delivery. When GBS gets into the lower urinary tract, it is a marker for heavier colonization. And actually in those individuals we don't even recommend culturing them again as they get closer to term. That bacterial colonization warrants empiric treatment of these folks during labor. And also if a mother has had a prior child affected with GBS disease, that is an additional indication for empirically treating her during labor with all of her subsequent pregnancies.

So having identified in the 70s that GBS was an important factor in neonatal sepsis, the early discussion was who should get screened during pregnancy? And that's where folks like you who work in the laboratory are so important. The initial debate was whether all pregnant people should be screened during pregnancy, or whether the identified risk factors should be the primary reasons that people got screened during pregnancy. It was identified in the 80s that giving folks who were colonized by culture targeted antibiotics during labor was a very effective way at preventing this early-onset disease in their newborns. This was identified in early studies in the 80s. And has been reinforced most recently by a systematic review in the journal Clinical Infectious Diseases about five years ago, showing that intrapartum antibiotics targeted against GBS is really the most effective way to prevent newborn illness. In terms of identifying colonized women and treating them intrapartum, that is really the most effective way to deliver antibiotics.

It has been shown that if you treat people, or culture them, before they go into labor and then try to treat them before labor or give oral or intramuscular regimens instead of intravenous antibiotics, none of those approaches has been shown to be effective. And then once it became established that colonized pregnant women should receive antibiotics during labor, the challenge was how to best identify these colonized women who should receive this prophylaxis.

When we use intrapartum antibiotics targeted against GBS, it's really based on a two-pronged approach. One is to lower the rates of neonatal GBS colonization. Because if a newborn isn't colonized, it won't get infected. And that requires that the drug used achieves adequate maternal drug levels to deal with colonization in the genitourinary tract. And also it needs to additionally reduce the risk of neonatal sepsis if a newborn is colonized. And that requires adequate levels of the antibiotic in the fetus and the newborn. So the current recommended antibiotics and dosages of antibiotics in the guidelines, were developed with the goals of both achieving adequate drug levels above a MIC for GBS in fetal blood and amniotic fluid, while minimizing the risk of maternal side effects and toxicity. So the very first iteration of the GBS guidelines were published by the CDC with input from the National Organizations for OB-GYNs, Pediatricians, and Microbiologists in 1996.

And it evaluated an approach and gave people the option of either screening everybody at 35 to 37 weeks, giving antibiotics for those who are identified as carriers. Or giving antibiotics for those who had preterm delivery. But also gave the option of risk-based approach and using antibiotics during labor only for people who developed risk factors. And in both strategies, prophylaxis was given to the women who we talked about earlier. Who had bacterial colonization during pregnancy, or who had a prior affected newborn. The problem, unfortunately for OBGYNs, is that they didn't really understand or remember what the risk factors were. And follow-up studies after this actually showed that the risk-based approach was clearly inferior to universal screening, primarily because providers missed a significantly greater number of at-risk women using a risk-based approach compared to screening. So this graph demonstrates the impact of subsequent guidelines after 1996 on the rate of early- and late-onset GBS.

And what you can see is that the initial guidelines were here in 1996. And as they started to become adopted, the rates of early-onset GBS declined. And then as subsequent iterations were rolled out, we have a significant drop in the rates of newborn GBS and late-onset GBS. Which is really a different disease, was not impacted. But that is not really what was intended. But really the era of universal screening rather than risk-based screening clearly made a difference in lowering these risks. So the most recent iteration of these guidelines, I had the opportunity to participate in being one of the editors for ACOG of these new guidelines. They were initially published in 2019, with a interim update and the final version in early 2020. And what we did is we changed a few things with folks like you in the lab will be seeing.

Initially we had recommended that GBS screening be done between 35 and 37 weeks gestation. And that's because studies suggested that the greatest predictive window for these pre-delivery GBS cultures was about five weeks. That was initially reported in the late 90s by the group at University of Florida using culture-based screening. And more recently by a group out of Finland, about three years ago looking at PCR-based pre-delivery screening. And both of those, using both culture and PCR-based screening, demonstrated a culture to delivery predictive window of four to five weeks.

And that that predictive window dropped significantly if the culture was done too much before delivery, certainly after five weeks. So when we evaluated the data for this new set of guidelines, we changed the screening window a bit to 36 to almost 38 weeks. And the reason for that is, if someone were to come in in labor prior to 36 weeks when they wouldn't normally be screened, the default would already be to treat them during labor because they were delivering prematurely. And this would provide a five-week window for a valid culture up to 41 weeks. And in the United States, there are many more women who deliver at or beyond 41 weeks, than those who deliver between 35 and 36 weeks. So we are actually getting more women falling within the valid predictive value when we move the screening window by just a week later, so that the cultures would continue to be valid up to around 41 weeks.

Part of the guidelines that directly impact laboratory personnel really have to do with how these tests are ordered and processed. We very strongly emphasize in the guidelines that when these vaginal-rectal swabs are being obtained, whether they're culture or whether they're PCR, they have to be marked as coming from a pregnant woman. And any penicillin allergy that that person has, needs to be noted on the laboratory request. Whether it's a paper requisition in a given institution, or more commonly in the electronic medical record. These swabs need to be done from more than one site that sampling only the vagina, for example, significantly lowers the sensitivity of culture and PCR by about 20%. So these need to be vaginal-rectal swabs. And recent studies have demonstrated that among providers incorrect sampling, typically vaginal only and not sampling the rectum, is the most common screening error. And we try to teach our residents this and we try to emphasize it when people like Dr. Pride and I give grad rounds. However, it takes a lot of repeated emphasis of how to do things properly.

The other aspect of processing is that the labs need to know that these are GBS screening swabs, so that these swabs need to be incubated first in a selective enrichment broth to optimize the sensitivity of results. And this applies both for culture and for PCR. And then if a woman reports a penicillin allergy that is significant enough to not use penicillin or a cephalosporin, then clindamycin susceptibility testing needs to be done for those individuals who report a penicillin allergy to help providers decide which alternate antibiotic would be used if the culture comes back positive. So we are entering with GBS screening the era of molecular testing. And those of you who work in the lab, know that molecular testing is not new for infectious agents. And it has evolved as a first line method for a number of infectious agents, specifically gonorrhea and chlamydia. Which, those of us who are my age, remember actually doing formal cultures for. But now we can do a duplex PCR, either on a swab, or in urine for both gonorrhea and chlamydia. And TB as well, is now primarily a molecular testing modality.

Nucleic acid amplification testing for GBS has been shown in a number of studies over and over again, to be at least equivalent but more commonly superior in sensitivity to culture-based screening when that test protocol includes a pre-analysis incubation step. And in a number of studies, one published about five years ago, shows that the sensitivity of culture versus PCR is culture is inferior to nucleic acids testing. And even though these numbers are close, they are still statistically significant. The caveat here is that rapid testing by PCR still carries with it with GBS a lower sensitivity. And that is why when we evaluated the data when we were writing these guidelines, we strongly advocated for the use of molecular-based testing as a reasonable and potentially more sensitive alternative to culture for antepartum screening, since the results needed to be available within six to 12 hours. And there would be time to optimize the sensitivity of the test by having the appropriate incubation phase before the aliquot in the analysis tube was actually put into the processor.

The other primary limitation with nucleic acid testing for GBS is for those individuals who report a severe penicillin allergy, because since PCR testing does not isolate an organism the way that culture does, you can't do susceptibility testing from the same swab or the same aliquot. And you need to be able to set up a separate culture to do susceptibility testing for clindamycin in folks who report a penicillin allergy. And that's why it's so critical for providers to report penicillin allergy to the lab if NAAT is being used for antepartum screening. We're still fighting to get PCR antepartum screening used routinely. The most recent CDC survey showed that while the laboratory use of NAAT-based GBS screening increased from 2010 to 2016, it is still low overall. 93% of the almost over 500 labs who were surveyed responded, which is impressive for laboratories to actually reply to that degree.

We can't get those kind of results when we screen medical groups unfortunately. And just under 20% actually reported using NAAT. Almost all the labs using NAAT were hospital or clinic-based. And almost all of them reported using the enrichment step, which means that they were using it for screening, not for rapid point of care testing on labor and delivery. And I'm happy to say that my new position in the division at UCLA, I get to work closely with our director of microbiology. And they use NAAT screening for antepartum screening. And that obviously makes me very happy that I don't have to try to militate for it in a new institution. And just why this is so important. This addresses finally sort of the issues that we have with antibiotic stewardship in the United States. We still keep penicillin as the first line agent for prophylaxis in folks who do not have severe allergies.

Ampicillin should still be reserved as an alternative. And penicillin is preferred because it has a narrower spectrum against gram-positive bacteria. And a lower risk of evolving resistance in other organisms that may be living in the vaginal flora. In a CDC surveillance study that was published eight years ago, it was demonstrated that while the rates of GBS neonatal sepsis continued to decrease even one or two doses of ampicillin as a non-necessary alternative for prophylaxis during labor, would cause the evolution of resistant E. coli to ampicillin in almost two thirds of exposed cases. And this really only required one to two doses during labor.

When folks report severe or high risk penicillin allergies meaning anaphylaxis, or more severe dermatologic manifestations, then the first line drug would be clindamycin. If this isolated specimen is susceptible to clindamycin, and if the GBS is clindamycin-resistant, then vancomycin is recommended. This is an important issue because almost 50% of GBS in the United States is now resistant to clindamycin. So that it's very important for us to have the susceptibility information so that we're not using clindamycin empirically with a resistant organism. And basically not treating the mom at all. And assessing penicillin allergy is clearly an important thing for us to be considering. And these most recent guidelines have emphasized the importance and safety of considering penicillin allergy testing during pregnancy, simply because the vast majority of people who report an allergy are actually tolerant at least 90% of the time. So I am going to wrap up that portion of my presentation, and I'm going to turn the floor over to Dr. Pride. Thank you for your attention.

David T. Pride MD, PhD:

Thank you so much for the introduction. It's my pleasure to be here today to talk to you all a bit about nucleic acid testing for Group B streptococcus. You guys have already seen Dr. Silverman provide a much more clinical and guidelines-based approach for Group B streptococcus testing. And I'll provide you guys the perspective of myself, who's an infectious diseases physician, as well as a director of a clinical microbiology laboratory. And I'm specifically at the University of California San Diego. So I'll start with my disclosures. First I am currently a consultant for Cepheid. I have been a consultant for both Roche and Hologic, who have some of the tests that we will speak a little bit about today. So a little bit about Group B streptococcus. As you guys have heard, it is a potential pathogen, but it's also a colonizer, or an organism that lives on the skin, of a lot of individuals.

And in general, it's an organism that doesn't really want to make you sick. And for a lot of pathogens that live on the human body, they tend to become pathogens when for example, they get to places where they're not supposed to be. So for example, with a common pathogen like staphylococcus, it lives on your skin and if you get a cut, or scrape, or scratch on your skin, then that pathogen can get underneath your skin and then cause disease. The same can be said of an organism like Group B streptococcus. It is the only sort of Lancefield Group B known as streptococcus, also known as streptococcus agalactiae. And it commonly lives on the skin. And maybe more important for today's topic is, it lives in the gastrointestinal tract as well as potentially on the vagina of women. And why that is a problem is not necessarily because it is typically very pathogenic in those women. But it's a problem in pregnant women because there's potential for transmission of the microbe to the relatively naive neonate.

So as I was just mentioning, we really are quite concerned about the potential for Group B streptococcus transmission. We tend to be concerned primarily during the process of childbirth, whereas that infant traverses the vaginal tract, it is able to pick up what may be colonizing Group B streptococcus. And then that infant ends up being born with Group B streptococcus, and then has the potential to develop a Group B streptococcus infection. The risk factors, I think you guys have kind of already heard about. The most obvious risk factor is the mother being colonized, but then there's the prolonged rupture of membranes. As well as premature deliveries that are risk factors for Group B streptococcus as well. And, as I sort of mentioned, it's not particularly pathogenic for most of us, as there are many of us, both men and women, who are colonized with the Group B streptococcus.

But where we worry about it most is in the transmission of those microbes to the neonates. According to the CDC, there's about 31,000 or so Group B streptococcus infections in the US every year. The majority of those, thank goodness, are adult infections. Where we worry most though, it causes about 7,600 infections in newborns every year. And it has about a 5% mortality rate. And while 7,600 may not sound like a lot of infants, when you think about a 5% mortality rate on that 7,600 infections, that's actually quite a few infants. We worry most about the early-onset disease that occurs generally within the first seven days of childbirth, but also there's a potential for later onset disease as well. So you could never... Or at least early on, you're not necessarily out of the clear just because that infant or neonate has not developed an infection by day seven.

And the sorts of diseases we may see in children who have become colonized as they are born, is that they may develop bacteremia, sepsis, pneumonia, as well as meningitis. And unfortunately the neonates are not very well-equipped to be able to deal with these sorts of infections. So while disease in mothers is not uncommon from Group B streptococcus, it typically is not something that is particularly fatal, for example. In young children, unfortunately, the case is a little bit different because they're not well-equipped to be able to deal with it. And the reason that that's so important is that Group B streptococcus is incredibly susceptible to antibiotics. We don't see a whole lot of organisms nowadays that are still susceptible, for example, to basic penicillin, which has been around for a very long period of time. Group B streptococcus remains highly susceptible to penicillin. However, sometimes you just can't deliver penicillin fast enough to be able to prevent morbidity and mortality in those neonates.

So as Dr. Silverman was saying, protocols have been developed for the treatment of Group B streptococcus. And the people to treat, I think to some extent, do become obvious, right? Group B streptococcus in the urine, as you might imagine, the vaginal tract, urinary tract are quite close in women. So if you find Group B streptococcus urinary tract infection, or just colonization of the urinary tract, really you need to treat for the potential that that neonate can become colonized. Any women with positive rectovaginal screens, meaning that you're screening what's living in the guts of those women, as well as what's living in the vaginal tract. If you find Group B strep, there's a possibility that that child can become colonized in the process of childbirth. If you don't know, in general, the recommendation is to treat. And there are still a number of women who still don't necessarily get routine prenatal care.

And in those circumstances, treatment really is the best option to try to prevent Group B streptococcus invasive disease. And then prolonged rupture of membranes is also a big risk factor. So you might imagine, with all of these protocols in place, why is Group B streptococcus still considered a problem? It seems like we're sort of perfectly prepared to be able to screen and reduce, as has been already shown, the prevalence of this particular disease. And the problem sort of becomes from a clinical microbiology standpoint, is in general, are we using the best test to be able to determine disease? What is the potential that we can be telling someone that they are negative, when in fact they are truly Group B streptococcus positive? And along with that might come risk to their infant? So the guidelines from the CDC really sort of recommend either Group B streptococcus culture, or Group B streptococcus nucleic acid amplification tests.

So in a lot of ways these tests are treated like their equivalent, and they're not exactly equivalent. It is true though that in the settings of a positive test and you treat, you can significantly decrease the prevalence of disease. But that doesn't necessarily mean that the tests that are available are equivalent in themselves. So there are culture-based tests as we sort of had mentioned. There are nucleic acid amplification-based tests that are also available. At my institution we did culture for a long period of time. And we somewhat recently have switched to nucleic acid amplification-based tests. And we were very happy with the culture that we were doing for many years. We had about a 20% positivity rate on our cultures. Primarily what we would do was we would take a rectovaginal swab as is customary for most Group B streptococcus testing. We take that rectovaginal swab, we do an enrichment step. Meaning that we take the swab, we place it in a media that will preferentially grow organisms like streptococcus, and then you take that enriched media and you try to grow it on a plate.

So that's what we did. And we would take advantage of the fact that Group B streptococcus is a beta hemolytic organism, so we could grow it on a blood agar plate and we could just look for evidence that there's lysis of the red blood cells on those blood agar plates. And that worked fairly well for us. There are also different types of media such as carrot broth, Granada agar, even Chromagar's, that allow you to do basically a similar thing to screen for patients who have Group B streptococcus infection. The problem in general, particularly with beta hemolysis is that somewhere around 4% to 5% of Group B streptococcus isolates are not beta hemolytic. And it's a problem as one might imagine, because if you imagine having a hundred pregnant women, all of whom have Group B streptococcus infection, and the best you might be able to do is to identify 95, 96 of them. Meaning that four or five are probably going to fall through the cracks.

And that's sort of the take that we have had on using culture for Group B streptococcus identification. That there's a small sort of window where you might be missing positives just based on the way some of the tests work. And it turns out that those same isolates that tend to test negative for beta hemolysis, also tend to test negative on carrot broth and Granada agar as well. So it's not really something that can simply be overcome just by changing the way you culture. And now there are nucleic acid amplification tests that are available which work quite well. But with that caveat that in general you're doing it after an enrichment step. So it's not the sort of test in general where you get an immediate answer. It's the sort of test where you take around a day to enrich, and then do a nucleic acid amplification test.

There are a lot of IVD or FDA approved assays for nucleic acid amplification of Group B streptococcus available. There's a test from Hologic, Luminex, there's several tests available from Cepheid. Even a rapid test without an enrichment step is available from Cepheid. And many of them do work similarly, or have a somewhat similar performance. But as we were sort of looking at transitioning from culture potentially to nucleic acid amplification-based test, we decided to do a rather large study really to evaluate how these tests may perform against one another. But most importantly to evaluate how they perform against culture. And we analyzed three different testing platforms. We looked at the Hologic Panther, a version of the Cepheid GeneXpert, as well as a version of the Luminex Aries. And we basically compared them using all of the same samples. And each of these different platforms has different capabilities and one might choose which of the platforms to use just based on the workflow of their own particular lab and the size of their institution, for example.

So at our institution, we do a lot of Group B streptococcus testing. So we really wanted to choose a platform that would allow us to do a relatively large number of tests within a day. So for example, you can see that there's the Hologic Panther, which can do a large number of tests. But there are different formulations of the Cepheid assays. Depending on which formulation you choose, you can do a relatively small number of tests or a larger number of tests. And the Luminex Aries as well has different formulations available that may allow you to do more or less tests. So those are the platforms basically that we analyzed in our analysis of our patient population. And we recruited 500 women, looked at 500 different vaginal-rectal swabs. Again, we subcultured them all using LIM broth. And then we took that LIM broth and we moved it to our standard of care, which is again culture on a blood agar plate looking specifically for Group B streptococcus.

And then we would identify that Group B streptococcus using multi-tall technology. And we compared those on the exact same specimens to different aliquots of that LIM broth tested on the Hologic platform, the Luminex platform, or the Cepheid GeneXpert platform. So in essence, we're comparing culture to nucleic acid amplification test. We're also comparing three separate nucleic acid amplification tests. And what we found, it wasn't terribly surprising, but it still to us was a little shocking when we actually looked at the numbers. We identified out of those 500, about a 108 of them, which were actually positive via culture. Again, right around 20% positive culture rate. And then when we look at those that were PCR positive or nucleic acid amplification tests positive, but culture negative, we could see substantial numbers using each of those tests. Which again was telling us, "Gee, maybe we've got more Group B streptococcus in our patient population than we had originally thought, based on our culture results."

So as we try to figure out, because we're looking at four different tests, which ones are right and which ones are wrong. We sort of decided on a really sort of simplistic consensus analysis to get at what we think are the true positives and the sort of false negatives. So in general, when we have four tests, in essence, if two or more are showing the same result, we would consider those true positives. And if, in essence, just one of the tests is showing us that it's positive, we might consider that a false positive. So in our analysis of culture, we found again, a 108 true positives, but 39 false negatives. And that corresponds to about a 73.5% sensitivity rate in our patient population. And this is similar to other numbers that you can see in the literature, which certainly strongly suggested that we're missing a number of Group B strep positives in our patient population.

When we look at the PCR test, you don't see nearly those numbers, false negatives. And in fact, each of them have a sensitivity right around 96%. And the number of false negatives is closer to five, or six, all the way across. So certainly using the sort of consensus criteria has given us the sense that these assays are more sensitive than is the culture-based technique. And when we saw that, we really sort of had to make an immediate decision that we've probably got to move on from culture. And one of the sort of anecdotes that I would say as to why that's really important to us, is that one of the things that people may not appreciate as much about life in a clinical lab. But for the most part in clinical labs, the people that work in clinical labs know, "This is bad." Or, "This is okay, we can deal with it."

And I can tell you in the clinical lab, every single time we see a neonate that's born and we get a Group B streptococcus culture that's positive from blood, or from the cerebral spinal fluid, or even from the lungs, everyone hearts sinks here. We realize the importance of it. And part of the reason that hearts sink over here, is that we realize that this is potentially preventable. If we're using highly sensitive tests to be able to screen mothers as best we can so that they could be prophylaxed properly against it, this is something that can be prevented. So that's sort of the way that we look at it in the clinical lab. So when we saw that number of 39, it was not something that we felt we could move forward with at our institution. When we look specifically at CT values, again, it's sort of the cycle threshold where a test becomes positive. The lower the CT value in general, the more reliable the result typically is going to be. And the more likely you are to have multiple different tests that give you the same sort of answer.

When we look at our true positives, the CT values are lower, somewhere around 20-ish. And for our false positives, they're much higher, closer to 40. And the one thing to keep in mind is just as a generality, CT values less than 30 are usually when there's just a lot of the organism around. CT values around 31 to 36 usually are more when there's a moderate amount of the organism that is around. So our CT values of around 20 or so, are telling us that there's a whole lot of this Group B streptococcus around in these patients where we're detecting it. And we can see that when we just look at concordance amongst our testing methods. When all four methods agreed with each other, generally the CT values were right at around 20, and in some cases below. Again, telling us that in order to get concordance, we have to have high amounts of the microbe present. When we see the three nucleic acid amplification tests that all agree with one another, in general, the CT values are right around 30.

Again telling us that we've got a relatively high to moderate amount of those microbes that are present. And that's sort of what has kind of concerned us, right? The idea that we could have high to moderate amounts of a microbe present and yet still miss it on culture. And that's what got us really to make the switch to Group B strep nucleic acid amplification test. And it's why we strongly suggest that others around the country in their laboratories make those switches as well. The one thing we are not absolutely certain of, is when you get a CT value of 37 or 35, how likely is that baby to be colonized, to become a problem? But we know that the risk is not zero when these tests are positive. So that we feel like moving to nucleic acid amplification assays are really going to be the best modality to help us stamp out disease as is best as possible.

So at our institution, we made the switch right around 2019. And before we had had a positivity rate on our Group B streptococcus cultures right around 18%, maybe a little bit higher than 18%. Since we moved to nucleic acid amplification test, our positivity rate's been at about 20% to 21%. So it's not, maybe as much as you might have expected from seeing the previous figures, but it's still a lot of patients. It really is. We do a fair number of screenings at our institution. It's a lot of people. And it does amount to potentially hundreds of missed Group B streptococcus calls that are not missed at our institution anymore. The other thing I do just want to bring up is that screening, as Dr. Silverman I think has told you guys earlier, is really, really important. And the one thing that really has come across, particularly at our institution, but really all across the world is 2020 when we were dealing with probably the biggest part of the Covid-19 or the SARS-CoV-2 pandemic, there were fewer screenings.

We could see it in our data. Everyone else is seeing it in their data. The positivity rate didn't really change at all. There were just less women who were being screened during that time. There were still plenty of pregnancies. It's just less women getting that sort of routine prenatal care. And that should sort of concern us all because again, this is a disease that is potentially preventable in most circumstances. So not getting screening can potentially lead to more Group B streptococcus disease. So with that, I'll just sort of summarize what our findings and take on Group B streptococcus is. One is that, of course we saw a dip in 2020. Thank goodness we've recovered, and I think we're back to sort of our baseline in terms of numbers of tests. The positivity rates, we definitely saw an increase of a percentage or two. Which actually at our institution corresponds to a relatively large number of patients who can be treated against Group B streptococcus.

We know that pretty much every Group B streptococcus culture we can detect with nucleic acid amplification test. And we had a number of Group B strep positive nucleic acid amplifications in our own institution that sort of really helped us make the decision to move away from culture and towards nucleic acid amplification. And for the most part, most of the nucleic acid amplification tests perform very similarly in our analysis with a similar number of potential of false positive results. So which platform you choose at your institution, I think in a large part, can really just be based on what works best for the workflow and for the patients at your institution. With that, I will sort of like to acknowledge the folks who've been involved in our studies. And turn it over for a Q&A session.