Isolation of known and potentially pathogenic tick-borne microorganisms from European ixodid ticks using tick cell lines

Ana M Palomar,Shonnette Premchand-Branker,Pilar Alberdi,Oxana A Belova,Anna Moniuszko-Malinowska,Olaf Kahl,Lesley Bell-Sakyi

doi:10.1016/j.ttbdis.2019.02.008

Abstract

Ticks harbour and, in many cases transmit to their vertebrate hosts, a wide variety of pathogenic, apathogenic and endosymbiotic microorganisms. Recent molecular analyses have greatly increased the range of bacterial species potentially associated with ticks, but in most cases cannot distinguish between surface contaminants, microorganisms present in the remains of the previous blood meal and truly intracellular or tissue-associated bacteria. Here we demonstrate how tick cell lines, primary cell cultures and organ cultures can be used to isolate and propagate bacteria from within embryonic and adult Ixodes ricinus, Dermacentor marginatus and Dermacentor reticulatus ticks originating from different parts of Europe. We isolated and partially characterised four new strains of Spiroplasma from The Netherlands, Spain and Poland, two new strains of Rickettsia raoultii from Russia and Poland, one strain of Rickettsia slovaca from Spain and a species of Mycobacterium from the UK. Comparison with published sequences showed that the Spiroplasma strains were closely related to Spiroplasma ixodetis and the Mycobacterium isolate belonged to the Mycobacterium chelonae complex, while the R. raoultii and R. slovaca strains were similar to previously-validated species.

Highlights

Ixodid ticks of the genera Ixodes and Dermacentor are the most widespread and important vector species infesting livestock and humans in Western, Northern, Central and Eastern Europe (Estrada-Peña et al, 2006; Medlock et al, 2013; Rubel et al, 2016)
36 primary embryo-derived cell cultures were screened for presence of bacteria (Table 3) as part of monitoring during attempted cell line establishment, a procedure that can take between one and Spiroplasma were detected in Giemsa-stained smears prepared from two primary cell cultures set up from embryonic I. ricinus from The Netherlands, 7 months after culture initiation
Successful in vitro isolation of tick-borne bacteria depends on having culture conditions suitable for supporting survival and growth of the microorganisms, whether they are intracellular or extracellular

Summary

Introduction

Ixodid ticks of the genera Ixodes and Dermacentor are the most widespread and important vector species infesting livestock and humans in Western, Northern, Central and Eastern Europe (Estrada-Peña et al, 2006; Medlock et al, 2013; Rubel et al, 2016). They transmit a broad range of viral, bacterial, protozoan and helminth pathogens of veterinary and/or medical importance (Heyman et al, 2010; Hubálek and Rudolf, 2012; Jongejan and Uilenberg, 2004; Otranto et al, 2013; Portillo et al, 2015; Socolovschi et al, 2009). PCR assays cannot distinguish between genomic DNA of viable and non-viable bacteria present in the sample, whereas only viable bacteria will grow in vitro

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Results

Conclusion