Leishmania major reaches distant cutaneous sites where it persists transiently while persisting durably in the primary dermal site and its draining lymph node: a study with laboratory mice - PubMed (original) (raw)

Leishmania major reaches distant cutaneous sites where it persists transiently while persisting durably in the primary dermal site and its draining lymph node: a study with laboratory mice

L Nicolas et al. Infect Immun. 2000 Dec.

Abstract

So far, studies of Leishmania persistence in mice have used injections of parasites administered either intravenously in the tail vein or subcutaneously in the footpad. These routes poorly reflect the natural conditions when the sandfly delivers metacyclic promastigotes intradermally. In this study B10D2 and BALB/c mice were inoculated within the ear dermis with 10(4) Leishmania major metacyclic promastigotes. The parasite load was monitored by quantitative PCR in different tissues from the dermal inoculation site to distant tissues. The two sites of multiplication and persistence of parasites were the site of L. major inoculation and the draining lymph node (DLN), with a different pattern in the two mouse inbred lines. These two organs were the only sites harboring parasites 12 months postinoculation, with the DLN of BALB/c mice harboring around 10(7) parasites, a stable load from months 3 to 12. In these two sites, 8 and 12 months after inoculation, interleukin 4 (IL-4), gamma interferon, and inducible nitric oxide synthase transcripts parallel the parasite load while IL-10 transcript levels remain high. In addition, at early time points until month 3, parasite DNA was also detected in distant tissues such as the contralateral noninoculated ear or the tail skin, indicating that blood was at least transiently disseminating the parasites. In contrast, L. major DNA in liver, spleen, and femoral bone marrow remained sporadic in mice of both lines. This study is discussed within the framework of Leishmania transmission from the vertebrate host to the sandfly vector, a complex process still poorly understood.

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Figures

FIG. 1

FIG. 1

Evolution of induration of the inoculated right ear for B10D2 or BALB/c mice after intradermal inoculation of 104 L. major (NIH 173) metacyclic promastigotes into the ear dermis. Measurements are expressed as the difference of thickness between inoculated ear and noninoculated ear. The data represent mean values ± standard deviations (error bars) (n = 10). ∗∗, after week 6, the inoculated ear of most BALB/c mice showed necrosis and loss of tissue and therefore the thickness of induration could not be measured further.

FIG. 2

FIG. 2

Standard curves of detection of kDNA of L. major in different tissue homogenates. Tenfold numbers of in vitro-grown L. major NIH 173 were spiked to tissue homogenates prior to DNA extraction and PCR. Data represent the means ± standard deviations (error bars) of at least two DNA extractions. kDNA amplicons were quantified by scanning agarose gels with Ampli Quant Windows software.

FIG. 3

FIG. 3

Monitoring of the parasitic load in B10D2 (black histograms) or BALB/c (white histograms) mice after intradermal inoculation of 104 L. major metacyclic promastigotes into the right ear dermis. Parasitic load is estimated by L. major kDNA PCR (see Fig. 2). Histograms represent mean values + standard deviations (error bars).

FIG. 4

FIG. 4

Quantitation by RT-PCR of mRNA transcribed from IL-4, IFN-γ, IL-10, and iNOS genes in ears, draining lymph nodes and distant skin (tail) of BALB/c or B10D2 mice. Tissues analyzed are right ear (RE), right retromaxillary lymph node (RLN), left ear (LE), left retromaxillary lymph node (LLN), and tail skin (TS) at month 8 (open circles) and RE and RLN at month 12 (black circles). Data of LE and LLN at month 12 were very similar to those of month 8 and are not shown on this figure. Circles represent individual mice, and bars indicate the means. The number of each transcript is normalized to 106 β-actin copies.

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