Differential infectivity of the Lyme disease spirochete Borrelia burgdorferi derived from Ixodes scapularis salivary glands and midgut.

Blood fed nymphal Ixodes scapularis Say infected with Borrelia burgdorferi were dissected to obtain salivary gland and midgut extracts. Extracts were inoculated into C3H/HeJ mice, and ear, heart, and bladder were cultured to determine comparative infectivity. Aliquots of extracts were then analyzed by quantitative polymerase chain reaction to determine the number of spirochetes inoculated into mice. A comparative median infectious dose (ID50) was determined for both salivary gland and midgut extract inoculations. Our data demonstrated a statistically significant difference (P < 0.002) in the ID50 derived from salivary gland (average = 18) versus midgut (average = 251) extracts needed to infect susceptible mice. A rationale for the differential infectivity of salivary and midgut derived spirochetes is discussed.

Genome size and organization in the blacklegged tick, Ixodes scapularis and the Southern cattle tick, Boophilus microplus.

Genome sizes and the organization of repetitive DNA were determined in the hard ticks Ixodes scapularis and Boophilus microplus using reassociation kinetics. The I. scapularis genome contains approximately 2.15 pg (2.1x10(3) Mbp) of DNA and consists of no foldback (FB), 27% highly repetitive (HR), 39% moderately repetitive (MR), and 34% unique DNA. The B. microplus genome contains 7.5 pg (7.1x10(3) Mbp) DNA, and consists of 0.82% FB, 31% HR, 38% MR, and 30% unique DNA. In both species, repetitive sequences occur in a mixture of long and short period interspersion but most (65-80%) of the DNA follows a pattern of short period interspersion. Genome size and organization in the three tick species so far examined are distinct from other arthropods in having a greater proportion of MR, a lower proportion of unique and HR DNA of very low sequence complexity.

Capillary feeding of specific dsRNA induces silencing of the isac gene in nymphal Ixodes scapularis ticks.

Ixodes scapularis transmits several pathogens including Borrelia burgdorferi. Bioactive compounds in tick saliva support tick feeding and influence pathogen transmission to the mammalian host. These studies utilized oral delivery of dsRNA to silence an anticomplement gene (isac) in I. scapularis nymphs. Silencing of isac significantly reduced fed-tick weight compared to delivery of control lacZ dsRNA, and immunoblots specific for FlaB protein indicated a reduction in spirochete load in isac-silenced infected nymphs. SDS-PAGE demonstrated that isac gene silencing affected expression of a number of salivary and non-salivary gland proteins in ticks. Finally, multiple isac cDNA homologues were cloned, and these may represent a new gene family coexpressed during tick feeding. This work presents a novel oral delivery approach for specific gene silencing in I. scapularis nymphs and characterizes the effect of isac on blood-feeding in an attempt to block transmission of B. burgdorferi.