NRL of vector-borne infections

The National Reference Laboratory performs routine and confirmatory diagnostics of mosquito-borne infections - West Nile fever, Dengue, Yellow fever, Chikungunya and Zika virus infections (PCR and serology), tick-borne infections - Crimean-Congo haemorrhagic fever (PCR and serology), Tick-borne encephalitis (PCR and serology) and Lyme disease (PCR and serology - routine diagnosis with ELISA and confirmatory with immunoblot), as well as diagnosis of hantavirus infections (haemorrhagic fever with renal syndrome), lymphocytic choriomeningitis, leptospiroses and listeriosis.

The laboratory organizes and conducts courses on the etiology, epidemiology, clinic, diagnosis and treatment of Lyme disease and other bacterial and viral vector-borne infections; leptospirosis and listeriosis, as well as individual training of graduate students, PhD students and microbiology specialists.

The NRL of Tickborne/Vector-borne Infections annually undergoes two external international quality control audits for the diagnostic work of Lyme borreliosis and West Nile fever conducted by the INSTAND Institute for Standardization and Documentation of Medical Laboratories, Germany. The laboratory participates in the National External Quality Control of Lyme Borreliosis Serological Diagnostics of the Laboratory Network in the country, conducted by the National Center for Lyme Disease Control.

he National Reference Laboratory for Tickborne/Vector-borne Infections, Listeria and Leptospires was established in 2007 by merging the NRL Leptospires, Borrelia and Listeria and the Arbovirus Laboratory. The NRL for Borelia, Leptospires and Listeria was founded in 1953 and recognized as a reference laboratory for leptospiroses since 1960. This period was associated with active research in the field of etiology, epidemiology and diagnosis of Lyme disease and viral hemorrhagic fevers.

The introduction of genetic techniques in tick, mosquito, rodent, and avian studies refines the detection of Lyme borreliosis vectors and reservoirs of major and minor importance, Crimean-Congo hemorrhagic fever (CCHF), West Nile fever (WNF), hantaviruses causing hemorrhagic fever with renal syndrome (HFRS), leptospirosis and human granulocytic anaplasmosis. The potential for co-infections in ticks and rodents is investigated. One of the first studies in Europe on the infectivity of Ixodes ricinus ticks with Borrelia and Anaplasma was carried out.

The major immunodominant proteins of Borrelia burgdorferi were cloned and expressed and their involvement in the antibody response in different phases of Lyme borreliosis was investigated. The immunodominant C6 peptide, and whole cell lysate applied as diagnostic antigens in ELISA were compared.

Circulating serotypes of pathogenic leptospires are being analyzed and specified. The introduction of molecular methods allows the early diagnosis of leptospirosis.

Innovative methods are applied and multiple molecular-genetic and serological systems were validated for the effective demonstration of vector-borne bacterial and viral pathogens as well as some zoonotic infections in clinical materials. As a result of improved diagnostic methods and approaches the first autochthonous cases of WNT in Bulgaria were detected and the independent evolution of the strain in the country was demonstrated. For the first time in Bulgaria and in the Eastern Balkan countries the circulation of Puumala hantavirus was demonstrated in both humans and rodents.

A phylogenetic analysis of Bulgarian CCHT strains from humans and ticks was performed showing their belonging to lineage 5 (Europe 1). For the first time, lineage 6 (Europe 2) was also demonstrated in ticks (now known as Aigai virus).

Large-scale pioneer serological studies on the prevalence of WNF, TBE, CCHF in the country were conducted. Healthy humans, patients with fever of unknown origin, and domestic animals studied. The results confirmed the circulation of those viruses and the expansion of WNV and Puumala hantavirus in South-Eastern Europe.

The accumulated potential enables a rapid and adequate response to infections posing a threat to public health, e.g. outbreaks of Zika, Lassa, and Ebola virus.

1. Christova I, Panayotova E, Groschup M, Trifonova I, Tchakarova S, Andrada Sas M. High seroprevalence for Crimean-Congo hemorrhagic fever virus in ruminants in the absence of reported human cases in many regions of Bulgaria. Exp Appl Acarol 2018, 75: 227-234 Q3, IF 760₍₂₀₁₇₎.
2. Christova I, Trifonova I, Panayotova E, Dimitrov H, Gladnishka. Molecular detection identification of Puumala orthohantavirus in Bulgaria. Infection, Genetics and Evolution. 2019, 70, 42-44. Q2, IF 2.611₍₂₀₁₈₎
3. Baymakova M, I. Christova, E. Panayotova, I. Trifonova, A. Chobanov, I. Daskalov, G. Popov, K. Plochev. West Nile virus infection with neurological disorders: A case report and a brief review of the situation in Bulgaria. Acta Clin. Croat. 2019; 58:546-549. Q3, IF 0.366₍₂₀₁₈₎
4. Christova I, Panayotova E, Trifonova I, Taseva E, Gladnishka T, Ivanova V. Serologic evidence of widespread Toscana virus infection in Bulgaria. J Infect Public Health 2020 Feb; 13(2):164-166 Q1, IF 2.487₍₂₀₁₈₎
5. Christova, I., A. Papa, I. Trifonova, E. Panayotova, S. Pappa, O. Mikov. West Nile virus lineage 2 in humans and mosquitoes in Bulgaria, 2018–2019. Journal of Clinical Virology, 2020, 127: 104365. Q2, IF 2,777
6. Pishmisheva-Peleva, M., Christova, I., Kotsev, St., Trifonova, I., Panayotova, E., Michaylova, R., Petrov, A., Popov, G. Stoicheva, M. West nile fever in Bulgaria - Clinical features - Analysis of a 2-year period. General Medicine, 2020, 22 (3), pp. 8-15. Q4, IF 0.04
7. Christova I, Trifonova I, Gladnishka T, Dragusheva E, Popov G, Ivanova V. Viral load and antibody levels in patients with COVID-19. GSC Advanced Research and Reviews 2021, 08(3), 10-18.
8. Christova I., I. Trifonova, E. Panayotova, H. Dimitrov, T. Gladnishka, V. Mitkovska, E. Taseva, I. Gergova, Hantaviruses in Small Mammals in Two Regions in Bulgaria. Acta Zoologica Bulgarica, 202173 (1), March 2021: 119-123. Q4, IF 0, 448 ₍₂₀₂₁₎
9. Nikolova M, Y. Todorova, R. Emilova, I. Trifonova, T. Gladnishka, N. Petrova-Yancheva, T. Cherveniakova, E. Dragusheva, G. Popov, I. Christova. Induction of humoral and cellular immune responses to COVID-19 mRNA and vector vaccines: A prospective cohort study in Bulgarian healthcare workers. J Med Virol 2022, 94, 2008-2018. Q1, IF 2, 327 ₍₂₀₂₀₎
10. Trifonova I, Christova I, Ivanova-Aleksandrova N. Gladnishka T, Ivanova V, Panayotova E, Taseva E, Dimitrov D, Marinov M, Kamenov G, Zehtindjiev P. Survey of Borrelia burgdorferisensu lato and West Nile fever virus in wild birds in Bulgaria. Biologia(2022) 2022 77(3):3519–3524, Q3, IF 1,65₍₂₀₂₁₎
11. Trifonova I, I. Christova, E. Panayotova, T. Gladnishka. RT-PCR systems for detection of arboviruses. Probl. Inf. Paras. Dis. 2018, 46 (1):25-28. Q4
12. Christova I., Trifonova I., Panayotova E., Tchakarova S., Gladnishka T., Taseva E., Seroprevalence study for West Nile virus in Bulgaria, Inf. Paras. Dis. 2018, 46 (1):29-31. Q4
13. Christova I, E. Panayotova, I. Trifonova, E. Taseva, T. Gladnishka. Unexpectedly high number of West Nile neuroinvasive diseases in Bulgaria in 2018. Probl. Inf. Paras. Dis. 2018, 46 (2):16-19. Q4
14. Taseva E, I. Christova, E. Panayotova, I. Trifonova. Investigations of murine rodents for the presence of leptospira DNA by nested PCR. Probl. Inf. Paras. Dis. 2018, 47 (1):25-29. Q4
15. Panayotova E, I. Christova, I. Trifonova, Taseva, T. Gladnishka, V. Ivanova. Seroprevalence of West Nile virus in Bulgaria, 2018. Probl. Inf. Parasit. Dis. 47, 2019, 2, 15-17. Q4
16. Taseva E, I. Christova, E. Panayotova, I. Trifonova, T. Gladnishka, V. Ivanova. Analysis of circulating strains, causing invasive listeriosis in Bulgaria, 2010-2019. Probl. Infect. Paras. Dis. 48, 2020, 2, 27-34.
17. Ivanova-Aleksandrova , I. Christova, D. Dimitrov, MP Marinov, Panayotova E, I. Trifonova, E. Taseva, T. Gladnishka, G. Kamenov, M. Ilieva, P. Zehtindjiev. Records on Ixodid ticks on wild birds in Bulgaria. Probl. Inf. Paras. Dis. 49, 2021, 2: 35-39.
18. Gladnishka T., I. Christova, I. Trifonova, V. Ivanova, E. Panayotova, E. Taseva. Study on ticks removed from patients for infection with Borrelia burgdorferi and their number depending on temperature and precipitation in 2016-2021. Probl. Inf. Paras. Dis.  2022, 50 (1): 32-36.