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Year : 2020  |  Volume : 57  |  Issue : 2  |  Page : 189-192

Kinetics of immune response to Francisella tularensis and Borrelia burgdorferi in a 10-year-old girl with oculoglandular form of tularemia after a tick bite: A case report

1 National Public Health Institute–National Institute of Hygiene, Warsaw, Poland
2 Public Provincial Specialist Hospital, Szczecin, Poland

Date of Submission24-Sep-2018
Date of Acceptance05-Apr-2019
Date of Web Publication14-Jul-2021

Correspondence Address:
Waldemar Rastawicki
National Public Health Institute–National Institute of Hygiene, 00–791 Warsaw, Chocimska 24
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/0972-9062.310870

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Keywords: Immune response; Lyme borreliosis; tularemia; tick bite

How to cite this article:
Rastawicki W, Chmielewski T, Łasecka-Zadrożna J. Kinetics of immune response to Francisella tularensis and Borrelia burgdorferi in a 10-year-old girl with oculoglandular form of tularemia after a tick bite: A case report. J Vector Borne Dis 2020;57:189-92

How to cite this URL:
Rastawicki W, Chmielewski T, Łasecka-Zadrożna J. Kinetics of immune response to Francisella tularensis and Borrelia burgdorferi in a 10-year-old girl with oculoglandular form of tularemia after a tick bite: A case report. J Vector Borne Dis [serial online] 2020 [cited 2023 Mar 30];57:189-92. Available from: http://www.jvbd.org//text.asp?2020/57/2/189/310870

Ticks transmit a great variety of pathogenic microorganisms and are the most important vectors of diseases affecting humans[1]. Furthermore, ticks can be infected with more than one pathogen and can simultaneously transmit various diseases[2]. Tularemia and Lyme borreliosis are among many different zoonotic diseases widespread in the northern hemisphere transmitted by ticks[3].

Tularemia is a highly contagious zoonotic disease caused by Francisella tularensis. Subtypes of tularemia include ulceroglandular, glandular, oculoglandular oropharyngeal, typhoidal, pneumonic, and gastrointestinal[4]. The ulceroglandular form is the most common, whereas the oculoglandular form of tularemia is one of the rarest, accounting for as few as 4% of all cases[5]. The latter form mostly occurs in the event of direct contamination of an eye; for example, when the eyes are rubbed after handling an infected animal carcass or when blood from a compressed tick is sprayed into an eye[6].

Lyme disease is a zoonotic tick-borne disease caused by spirochete bacteria belonging to the genus Borrelia. Clinical manifestations in humans include erythema migrans in the early phase, followed by neuro-borreliosis, Lyme arthritis and/or Borrelia lymphocytoma several weeks later. Similar to tularemia, Ixodes ricinus is the most common tick responsible for transmission of Lyme disease. Transmission of Borrelia occurs through injection of tick saliva during feeding[7].

Tularemia as well as Lyme borreliosis can be difficult to diagnose, especially in the early stages. They are relatively rare diseases, and their symptoms can be similar to other more common illnesses. Routine laboratory diagnosis of both diseases is mainly based on serological investigation. Antibodies to the pathogens appear in infected individuals, usually a few days or weeks after the onset of clinical symptoms and may persist at a diagnostically significant level for several months or even years[7],[8],[9].

In the present study, we describe the clinical case of a child with the oculoglandular form of tularemia also infected by Borrelia burgdorferi sensu lato after a tick bite. To analyze the kinetics of immune response in the course of tularemia and Lyme disease, we determined the level of antibodies to F. tularensis and B. burgdorferi in serum samples obtained at various intervals during a follow-up period, extending two weeks to one year after the onset of the clinical symptoms of the diseases.

Case report

A 10-yr-old girl was admitted in June 2016 to the Children’s Observation and Infectious Ward of the Provincial Specialist Hospital in Szczecin, Poland due to fever, swollen eyelid of the right eye, erythema of the right cheek and conjunctivitis. According to the anamneses, one week before admission to the hospital, during the child’s stay at a camp in the forest, symptoms of fever and vomiting suggesting angina had appeared. The girl was treated with amoxicillin for 5 days without any results. At this time, a tick was removed from the skin of the upper right eyelid. The exact date of the tick bite is unknown. There had been no other tick bites in this child in the past. Physical examination showed significant swelling of the right eye eyelid, with red skin, conjunctivitis with abundant purulent content, without obviously obstructed nasal patency and without symptoms of meningitis. In additional tests, the inflammation markers were slightly increased (CRP 80.9 mg/L), and the remaining test results were normal. Due to the notification of the tick bite the possibility of Lyme disease was taken into account, and a blood sample was taken for serological examination. The serological tests carried out using ELISA in the hospital revealed the presence of IgG and IgM antibodies to B. burgdorferi in the serum sample. Positive results were confirmed by Western-blot. However, despite the use of broad-spectrum antibiotic therapy, fever did not decrease and local symptoms were not resolved. In laboratory tests, an increase in inflammation rates was noted, and locally, a tender reaction from the pre-paramedomic lymph nodes and submandibular nodes on the right side appeared. A complex etiology of inflammation was suspected, with the possibility of co-infection with other pathogens. On the sixth day of the hospital stay, doxycycline was administered. Treatment resulted in a return to normal body temperature, improvement in local changes and the levels of inflammation markers decreased. The patient remained asymptomatic and was discharged to return home. After the discharge of the child from the hospital, the results of tests confirming tularemia were obtained. The girl was examined several times on control visits in the Children’s Observation and Infectious Ward Outpatient Clinic. She was in a good condition, without any symptoms signaling a recurrence of the inflammatory process.

The material for the study included five serum samples, taken at: 2 weeks (sample no. 1), 6 weeks (sample no. 2), 3 months (sample no. 3), 6 months (sample no. 4) and 12 months (sample no. 5) after the onset of the first clinical symptoms. The in-house ELISA was designed to detect anti-F. tularensis IgA, IgG and IgM antibodies in serum samples. The test was carried out on Nunc Maxi-Sorp polystyrene plates, according to the method previously described[8]. B. burgdorferi IgM and IgG antibodies were tested with the ELISA test (EIA Borrelia 14kDa+OspC IgM and EIA Borrelia IgG+VlsE, DRG-Medtek, Germany). All results were calculated as a cut-off index (COI) and interpreted as positive when the range was >1.1. Positive results were confirmed by the Western-blot method (Euroline Borrelia-RN-AT IgM, Euroline Borrelia-RN-AT IgG; EUROIMMUN, Lubeck, Germany). Results were interpreted as seropositive according to criteria of the German Society of Hygiene and Microbiology.

[Figure 1] shows the antibody levels for F. tularensis, expressed as OD values at 450nm, in five serum samples diluted 1/3200, collected during one year. In viewing the antibody decay profiles of the antibody response, it is clear that IgA, IgG and IgM antibodies can be detected in high levels in all serum samples. In the first serum sample, obtained approximately 2 weeks after the onset of symptoms, the IgA, IgG and IgM antibody level was approximately 0.2. In the second serum sample, obtained after another 1 month, the IgA class antibodies increased sharply to about 1.0, IgM class antibodies to 1.3, while the IgG class antibodies to OD450 nm to about 0.75. Examination of the third serum sample showed a further increase in the level of antibodies in all three classes of immunoglobulins, with the highest levels achieved by IgA. In the fourth serum sample, obtained approximately 6 months after the onset of the clinical symptoms, a small but consistent increase in IgA, persistence of IgG and a clear decrease in IgM class antibodies was observed. A slight decrease in the level of IgA and IgG antibodies as well as a further, sharp decrease in IgM class antibodies was observed in the fifth serum sample.
Figure 1: Levels of antibodies to F. tularensis determined by in-house ELISA (OD450) in five serum samples diluted 1/3200, collected from a 10-yr-old child with a oculoglandular tularemia during one year at different times from the onset of clinical symptoms. Diagnostic cut-off values for IgA, IgG and IgM antibodies are below 0.1.

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The serological investigations also showed that in all five tested serum samples specific antibodies to B. burgdorferi were detected [Figure 2]. The IgM response against spirochetes in consecutive serum samples showed a gradual decrease from 2.1 COI to 1.5 COI, starting within the first month after the treatment with amoxicillin and doxycycline. A significant increase in the IgG antibodies from 1.8 to 2.4 COI was seen between 2 and 6 weeks of follow-up. After that time, the patient had recovered completely and the next sera showed a gradually decreasing IgG level at 12 months of follow-up to the value of 1.6 COI. The positive results from ELISA were confirmed by Western-blot. Specific antibodies to p83, p41, p39, OspC and LipidBa antigens in IgG and p41 and OspC in IgM were detected. The intensity of the staining of the bands did not differ in the individual samples.
Figure 2: Levels of antibodies to B. burgdorferi determined by ELISA (cut-off index) in five serum samples collected from a 10-yr-old child with a oculoglandular tularemia during one year at different times from the onset of clinical symptoms. Diagnostic cut-off value for IgM and IgG antibodies is 1.0.

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Ethical statement

The study was pursued following the approval of the Institutional Review Board (IRB) at the National Institute of Public Heath – National Institute of Hygiene, issued on 28.06.2012, Approval No: 4/2012.

  Discussion Top

Laboratory diagnostics of tularemia is based on confirmation of the presence of microorganisms or their DNA in samples of clinical material or, more often, on the detection of specific antibodies at diagnostic levels in serum samples. Serology is a useful method for routine diagnosis of tularemia because, during F. tularensis infections, there is strong, immunospecific and long-lasting humoral immunity[9],[10],[11],[12]. Generally, during most bacterial infections, IgM and IgA antibodies are connected with primary immune response and decline within some months after the onset of symptoms whereas high titers of IgG are characteristic for secondary immune response and reach a peak in the later phase of the disease. However, F. tularensis seems to be exceptional among infectious diseases as it induces a pattern of humoral immunity which includes continued synthesis of all three classes of antibodies for years after infection[9]. Confirmation of this can be seen in the results of our study in which a very high level of all three classes of immunoglobulin to F. tularensis was shown one year after the onset of infection. It is interesting that in this late stage we can see a higher level of IgA antibodies than IgG antibodies, which are generally considered to persist much longer than IgA. The mechanism stimulating the continuous formation of these antibodies may be connected with the intracellular character of F. tularensis[12].

As demonstrated by the graphs, the IgM and IgG antibodies to B. burgdorferi seemed to rise and decline slowly or remain stationary for months after therapy despite clinical recovery. The long interval between pretreatment and post-treatment samples may give the false impression of a stable value even though there might have been an increase in the OD in the intervening period. Earlier studies have demonstrated that more than 40% of patients were still positive in terms of IgM at one-year post baseline and more than 80% patients whose initial serum specimens were positive for IgG antibodies according to ELISA had positive titers of IgG at follow-up at over one year[15]. Most cases showed a consistent number, type, and intensity of IgM and IgG bands in Western-blot over the entire follow-up period[13],[14],[15]. Thus, this method may not be useful for monitoring the response to treatment of Lyme borreliosis. Such a phenomenon was also observed in our described mixed infection case.

It is known that co-infection of humans with different vector borne diseases may be a result of a single tick bite by an arthropod infected with several pathogens, or as a result of multiple bites by ticks infected with one pathogen[16]. The epidemiological data showed that such co-presence of vector borne diseases caused by different pathogens is common in humans and animals.

Frequent co-presence of antibodies to B. burgdorferi, A. phagocytophilum, Bartonella spp. and B. microti has been found in hunters, forest workers and farmers in Poland[16],[17]. Also, we have previously described cases of mixed infections by F. tularensis and Rickettsia spp.[18]. Co-infection of F. tularensis with B. burgdorferi was identified in 6% of black rats in Bulgaria[19]. Studies on Lyme borreliosis and other tick-borne zoonoses in the Austrian and Slovakian border area revealed a relatively high prevalence of co-infection with B. burgdorferi s.l. and F. tularensis in small terrestrial mammals, as well as in ticks[20]. Co-infection by B. burgdorferi sensu lato and F. tularensis appeared in 8.4% of ticks in Serbia[21].

In the available literature, there is only one report describing a case of dual infection, tularemia and Lyme borreliosis, acquired by a single tick bite in northwest Croatia[22]. Unfortunately, in the case report presented by us, the tick removed from the eyelid of the patient was not collected for further research by PCR method. Thus, there is no direct evidence that both the tularemia and Lyme disease occurred after a single tick bite. However, because there had not been any other tick bites in this child in the past, such a probability could not be excluded. Awareness of this phenomenon of co-infections may be important in the diagnosis of tick-borne disease, particularly given that the clinical picture and course of disease may be difficult to interpret.

The golden rule in serodiagnosis of many infectious diseases is monitoring of antibody titers in serum during acute and convalescent phases. However, it should be borne in mind that long-lasting humoral immunity as in tularemia can create problems in some clinical situations. Detection of antibodies to previous F. tularensis infection may be the reason for the discontinuation of the diagnosis of other etiological agents that cause diseases with a similar clinical picture to tularemia. Furthermore, the same diagnostic situation may be noted in the case of simultaneous co-infections. The case report described in our study best demonstrates that a complex diagnostic examination after a tick bite should cover various etiological factors.

  References Top

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  [Figure 1], [Figure 2]

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