BACKGROUND: Lyme disease, caused by Borrelia burgdorferi sensu lato (s.l.), is the most common vector-borne disease in the Northern Hemisphere, with Ixodes ticks as its primary vectors. However, many patients do not recall tick bites, fueling speculation about alternative transmission routes, particularly via mosquito bites. This belief is reinforced by studies reporting Borrelia presence in mosquitoes. This study evaluates whether three mosquito species can acquire, maintain, and transmit Borrelia spirochetes. METHODS: Mosquitoes (Aedes aegypti, Culex quinquefasciatus, and Culex pipiens biotype molestus) were fed on Borrelia-infected mice to assess pathogen acquisition. Additional experiments involved ex vivo feeding on Borrelia-enriched blood to examine spirochete persistence in the mosquito gut. The potential for mechanical transmission was tested by simulating interrupted feeding between infected and naive hosts. The role of trypsin in Borrelia survival and infectivity was also investigated. RESULTS: Mosquitoes exhibited low efficiency in acquiring Borrelia from infected hosts. Spirochetes artificially introduced through ex vivo blood meals were rapidly eliminated during digestion, primarily due to trypsin activity. Inhibition of trypsin prolonged spirochete persistence and infectivity in the mosquito gut. Mechanical transmission experiments revealed no evidence of Borrelia transmission from infected to naive hosts. CONCLUSIONS: Our findings demonstrate that mosquitoes lack the biological capacity to efficiently acquire and maintain B. burgdorferi s.l. spirochetes and are unable to transmit them through natural or mechanical means. This study provides compelling evidence against mosquito-borne transmission of Lyme disease and reinforces Ixodes ticks as the sole competent vectors, which is crucial for targeted public health interventions and accurate risk communication.

• Keywords
• Borrelia, Borreliosis, Lyme disease, Mosquito, Tick, Transmission,
• MeSH
• Aedes * microbiology   MeSH
• Borrelia burgdorferi * physiology   MeSH
• Culex * microbiology   MeSH
• Ixodes microbiology   MeSH
• Mosquito Vectors * microbiology   MeSH
• Lyme Disease * transmission   microbiology   MeSH
• Mice MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

Spirochetal bacteria were successfully isolated from mosquitoes (Culex pipiens, Aedes cinereus) in the Czech Republic between 1999 and 2002. Preliminary 16S rRNA phylogenetic sequence analysis showed that these strains differed significantly from other spirochetal genera within the family Spirochaetaceae and suggested a novel bacterial genus in this family. To obtain more comprehensive genomic information of these isolates, we used Illumina MiSeq and Oxford Nanopore technologies to sequence four genomes of these spirochetes (BR151, BR149, BR193, BR208). The overall size of the genomes varied between 1.68 and 1.78 Mb; the GC content ranged from 38.5 to 45.8%. Draft genomes were compared to 36 publicly available genomes encompassing eight genera from the class Spirochaetes. A phylogeny generated from orthologous genes across all taxa and the percentage of conserved proteins (POCP) confirmed the genus status of these novel spirochetes. The genus Entomospira gen. nov. is proposed with BR151 selected as type species of the genus. For this isolate and the closest related isolate, BR149, we propose the species name Entomospira culicis sp. nov. The two other isolates BR208 and BR193 are named Entomospira nematocera sp. nov. (BR208) and Entomospira entomophilus sp. nov. (BR193). Finally, we discuss their interesting phylogenetic positioning.

• MeSH
• Arthropods genetics   MeSH
• DNA, Bacterial genetics   MeSH
• Phylogeny MeSH
• RNA, Ribosomal, 16S genetics   MeSH
• Sequence Analysis, DNA methods   MeSH
• Spirochaeta genetics   MeSH
• Spirochaetales classification   genetics   isolation & purification   MeSH
• Bacterial Typing Techniques methods   MeSH
• Base Composition genetics   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DNA, Bacterial MeSH
• RNA, Ribosomal, 16S MeSH

We have screened 91 migratory birds representing 32 species during the autumn of 2003 for the presence of the zoonotic pathogens Borrelia and Chlamydophila. Using polymerase chain reaction (PCR), B. burgdorferi sensu stricto was detected in cloacal swabs and, in two causes, also in throat swabs in 8 individuals (8.7 %) representing 7 birds species; B. garinii and B. afzelii were not detected. C. psittaci was detected only in cloacal swabs; 6 birds (6.6 %) from four species were found to be positive. The PCR products were sequenced and the sequences were compared phylogenetically with the gene sequences of 14 Chlamydophila strains retrieved from nucleotide databases; although the sequenced DNA was only 110 bp long, all obtained sequences created a new cluster with sublines branching from a position close to the periphery of the genus. All tested samples appear distinct within the known species and were most similar to C. felis or C. abortis.

• MeSH
• Borrelia burgdorferi classification   genetics   isolation & purification   MeSH
• Chlamydophila psittaci classification   genetics   isolation & purification   MeSH
• DNA, Bacterial analysis   chemistry   genetics   MeSH
• Pharynx microbiology   MeSH
• Phylogeny MeSH
• Cloaca microbiology   MeSH
• Lyme Disease epidemiology   microbiology   veterinary   MeSH
• Bird Diseases epidemiology   microbiology   MeSH
• Polymerase Chain Reaction MeSH
• Psittacosis epidemiology   microbiology   veterinary   MeSH
• Birds microbiology   MeSH
• DNA, Ribosomal chemistry   genetics   MeSH
• RNA, Ribosomal, 16S genetics   MeSH
• Sequence Homology, Nucleic Acid MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Slovakia epidemiology   MeSH
• Names of Substances
• DNA, Bacterial MeSH
• DNA, Ribosomal MeSH
• RNA, Ribosomal, 16S MeSH