Tag Archives: Dr. Mayla Hsu

Lyme Arthritis: The Antibody Connection

by Mayla Hsu, Ph.D., Director of Research and Science, GLA

Lyme disease, caused by Borrelia burgdorferi bacteria, often leads to arthritis, with up to 60 percent of late-stage patients suffering from painful joints and swelling. Symptoms can persist for months and even years. Its suspected that a protein called arthritis-related protein (Arp), which is located on the surface of the Lyme bacteria causes these symptoms. Mice experimentally infected with B. burgdorferi develop arthritis symptoms that are resolved when the animals begin to make anti-Arp antibodies.

But an enduring medical mystery has been why these anti-Arp antibodies coincide with resolving Lyme arthritis but not any other aspect of Lyme disease. One reason could be that other proteins, also on the bacterial surface, may be protecting Arp and the bacteria from antibodies. And if those shielding proteins constantly adapt, they could be even more effective at protecting Arp.

It turns out that this shielding mechanism might be possible. Recent data reported by Abdul Lone and Troy Bankhead at Washington State University indicates that another bacterial surface protein known as  VlsE, may protect the bacteria from anti-Arp antibodies and act as a shield to prevent the immune system from fighting Lyme.

The mechanism of how VlsE defends the Lyme bacteria from Arp antibodies is an interesting story. Earlier work by this research team tested a mutant B. burgdorferi that lacks VlsE protein. This mutant was tested for its ability to infect and grow in immunodeficient mice. Growing the bacteria in mice (in vivo) is more informative than growing the bacteria in a test tube (in vitro), because a mouse has the complexity of an animal with multiple organ systems, and the bacteria disseminate through the body that has different cell types, a circulatory system and other features not found in vitro. Immunodeficient mice lack a functioning immune system, so growing bacteria in them allows observation of how the bacteria replicate unimpeded, in a complex animal system. 

In severe combined immunodeficient (SCID) mice, the VlsE-deficient mouse was capable of infection, showing no growth deficiency. However, if the animals were treated with anti-Arp antibodies, the animals were able to clear the infection.  This suggests that anti-Arp antibodies can stick to Arp on the bacteria and promote their destruction. In contrast, normal bacteria that have VlsE on their surface could not be cleared by anti-Arp antibodies in SCID mice. The contrast in bacterial clearance in the mice shows that VlsE somehow affects the clearance of bacteria by anti-Arp antibodies.

Next, the investigative team used microscopy to observe whether anti-Arp antibodies can stick to B. burgdorferi spirochetes when VlsE is also present on the bacterial surface. They found that although anti-Arp antibodies can bind to VlsE-negative bacteria, they cannot do so in VlsE-positive bacteria. This result suggests that VlsE may actually be a physical barrier that prevents anti-Arp antibodies from sticking to their target, the Arp protein. Precisely how VlsE does this is an interesting question. For example, are Arp and VlsE close together on the bacterial surface? Is VlsE larger or does it block only the specific epitopes, or subdomains that are bound by Arp antibodies?

Moreover, there were limits on VlsE-mediated protection of B. burgdorferi proteins. Control experiments indicated that VlsE may not block antibodies other than those targeting Arp. This was shown by using a blood serum (antisera) purified from mice infected with an Arp-negative, VlsE-negative bacteria. These antisera would lack antibodies against Arp and VlsE, but have antibodies against other B. burgdorferi proteins. Mice were pretreated with the Arp-negative, VlsE-negative antisera and then challenged with normal bacteria which had both Arp and VlsE. The normal bacteria could not infect, which showed that the presence of VlsE could not protect the bacteria from antibodies that target non-Arp bacterial antigens. This suggests that VlsE may only protect Arp, and not other Borrelia antigens.

Work by other investigators has shown that VlsE undergoes extensive antigen variation. The antibodies elicited by VlsE also change during disease course. So, the work of Lone and Bankhead raises questions about whether specific VlsE variants protect Arp, and if there is a correlation with disease stage and the onset of arthritis.

These findings are also important because they further highlight how, in addition to other immune dysfunctions, the bacteria have evolved multiple ways to specifically evade the humoral, or antibody-mediated immune response. Not only does the VlsE protein change during bacterial infection, providing a mechanism of immune evasion and persistence for the bacteria, but it may also protect other bacterial proteins from being targeted. Further studies will clarify how VlsE shields Arp.

And it is another piece of the puzzle of how B. burgdorferi may cause persistent arthritis. Previous findings implicated peptidoglycan, a different component of the bacterial cell wall. How these pieces fit together will explain disease outcomes and long-term symptoms.

Related Blogs:

Research POV: Lyme Arthritis and Peptidoglycan
Possible Clue to Lyme Arthritis Found in People’s Inflamed Joints


Video Interview with Dr. Neeta Connally at WCSU

Dr. Mayla Hsu, Ph.D., Director of Research and Science at Global Lyme Alliance interviews Dr. Neeta Connally, Ph.D., medical entomologist, associate professor and Director of the Tickborne Disease Prevention Laboratory at Western Connecticut State University.

Dr. Neeta Connally’s research efforts are aimed at the prevention of Lyme and other tick-borne diseases primarily those transmitted by the black-legged tick in the northeastern United States. In particular Dr. Connally’s lab and field projects focus on prevention in the backyard, interfering with the tick life cycle, or by changing patterns of human behavior.

Watch the educational video interview below:

Read a portion of the transcript below:

Dr. Hsu: Good morning. I’m Mayla Hsu, Director of Research and Science at Global Lyme Alliance. I’m here today at Western Connecticut State University with Dr. Neeta Connally, who is a professor of biology here at the university. Neeta, glad to meet you.

Dr. Connally: Great to see you.

Dr. Hsu: So you’re trained as a medical entomologist. Do you want to tell me what a PhD in medical entomology does?

Dr. Connally: Absolutely. A medical entomologist is someone who studies insects and arthropods that transmit diseases to humans, and so that can include such organisms as mosquitoes, or ticks, or sandflies, and fleas, and so really we know that a lot of insects and arthropods will carry pathogens and disease-causing organisms, bacteria, viruses, parasites, and they can transmit those to humans.

Dr. Hsu: So your major research interest is ticks, isn’t it? Do you want to tell me a little bit about your work?

Dr. Connally: The primary focus of the research program here in my lab is the prevention of tickborne diseases in the Northeastern United States, primarily those transmitted by the black-legged tick, also known as the deer tick. Our focus is on prevention, really zoomed into the landscape of one’s own backyard, and we know that a lot of people in this region of the country are exposed to ticks in their own backyards, so we’re trying to do a better job at understanding the science behind how we can keep people from getting a tick bite and getting a tickborne illness.

Dr. Hsu: Okay, so your main focus sounds like it’s on ticks. As we all know, ticks are carriers of Lyme disease and other bacterial and viral diseases. Do you want to tell me more about the diseases that are spread by these ticks?

Dr. Connally: Oh sure. In our region, which is here in the Northeastern US, the primary tick that bites humans is the black-legged tick, or the deer tick, and we know that tick to be implicated in five primary infectious agents of humans. Those five major ones are of course Lyme disease, and the one that causes human granulocytic anaplasmosis, which has been called ehrlichiosis. Some people still call it that. We have the babesiosis, which is a red blood cell parasite, that’s very much looks like malaria, and then we have a viral agent that is known as … It’s known as Powassan virus, but this variant of this disease, or of this virus, has been called deer tick virus, so then we also have a newly identified borrelia, sort of looks a little like our Lyme disease bacteria. It’s called Borrelia miyamotoi, and that causes an illness that can have sort of a relapsing fever type of clinical presentation.

Dr. Connally: So those are the five major ones associated with this tick, and you know, the tick is full of all sorts of microorganisms, so I think that we know there’s a lot of things in the tick, so I think that only time will tell what, if any role those other microorganisms play. For example, that Borrelia miyamotoi, when I was a PhD student, we knew that it was there in some of our ticks, but we didn’t think it was a human pathogen, so we sort of discounted it and didn’t think anything of it. It wasn’t until very recently that we know that it does cause disease. So I think we know there’s a lot of things in these ticks, so right now, we say these are the five major ones. Doesn’t mean that there aren’t other things in those ticks that are causing people to be sick.

Dr. Hsu: Now, what’s particularly special about your work, that I understand you received a very large research grant from the Centers for Disease Control very recently, to set up a very kind of unique study?

Dr. Connally: Yeah. We’ve been really fortunate to partner with the Centers for Disease Control for many years, and work on various types of intervention studies, looking at ways we could reduce ticks, and if that tick reduction will result in the reduction of people being bitten by ticks, and also in the reduction of the incidence of Lyme disease and other tickborne illnesses. So in 2016, we received a CDC grant to conduct what we call the Backyard-integrated Tick Management Study. In this study, we’re looking at a two-pronged approach to controlling ticks in the backyard, and we’re also looking at a neighborhood sort of approach versus just treating your own house, so we’re trying to understand if you and your neighbors got together and tried to kill ticks in your backyards, does that have a bigger impact than just you doing it alone in your own backyard? So we’re implementing a tick spray one time a year, and then we implement a rodent target bait box to kill ticks on the small mammals that carry the Lyme bacteria.

Dr. Connally: So yes, it’s a three-year study, and we implement the tick control for two years, but we also follow the people in the household, so we’re trying to understand what they do outdoors, how they use their backyard landscape, and also how often they’re exposed to ticks, and how often they will get a tickborne illness. So it’s a very multifaceted, intensive study.

Dr. Hsu:  I think one of the things that struck me, that was really strong about your study, is that you’re also incorporating a placebo control into it, right? So some of the yards get a pesticide spray and some of the yards don’t. And I think you told me once that people don’t know.

Dr. Hsu:  Why is that a strength of your study?

Dr. Connally: Absolutely. Because we have humans enrolled in the study, and we want to know what happens to them, it’s very important that they don’t know if they have received treatment. So what we do is we implement either the true treatment, or we implement a placebo, and it’s a blinded study, which means that the homeowners and their families who are enrolled know they’re getting something put in their yard, but they don’t know if it’s the true treatment or if they’re getting the placebo, and the placebo would be instead of the pesticide, we spray water, and instead of a bait box that contains the treatment for the ticks in it, it doesn’t have that. It’s an empty bait box.

Dr. Hsu: So that’s supposed to control for their behavior, if they think that they … If they don’t know if they have active pesticide or active bait boxes, they’re not going to alter their behavior?

Dr. Connally: Exactly. Right. We don’t want them to feel a sense of security and therefore change what other preventive measures they may take, so this is … Using placebos is very classic in many other types of study designs, and it makes sense to do it in this case, because we really need to understand the human outcomes. And if we were just measuring ticks, we could do that without a placebo, but because we want to measure what the humans are doing, we need to have them not know.

Dr. Hsu: And if I understand, you also have students, then, going and checking the yards, dragging the yards on a systematic basis, to count how many ticks they have. How often do the students go?

Dr. Connally: Oh yeah. So, we have student interns, who get trained in tick collection, and they help us go out and collect ticks from the properties of the homes, all of the homes enrolled in our study. And our study is taking place here in Western Connecticut and also in Southern Rhode Island with our collaborator Tom Mather, at the University of Rhode Island, so there are students from URI as well, who participate in this. So the students go out during the nymph season of the tick. That’s the poppy seed sized tick stage, that is most implicated in causing Lyme disease, just because of the time of year that it’s active, and it’s very difficult to see. So we are collecting nymph-stage ticks between late May and July. We go out multiple times to each of the properties, and we collect ticks from all those properties, and we can then assess the density of ticks in those backyards, and we can also make sure those are indeed the black-legged tick or the deer tick, and not some other tick species.

For the full interview, watch the video above.