Metacyclogenesis is a major determinant of Leishmania promastigote virulence and attenuation

Abstract

The in vivo virulence patterns of promastigote populations defined on the basis of agglutination by the lectin peanut agglutinin (PNA) were studied for various cloned lines of Leishmania major. Promastigotes derived from logarithmic-phase cultures, which were routinely 100% agglutinated at 100 micrograms of PNA per ml, were relatively avirulent for BALB/c mice. The relative virulence of stationary-phase promastigotes appeared to be attributable to the proportion of nonagglutinable (PNA-) promastigotes contained within these populations. Purification of PNA- organisms from stationary cultures provided for each clone the most virulent inoculum, supporting the view that this change in lectin binding accurately reflects the development of infective metacyclic stage promastigotes. By studying this marker, we found that there was considerable variation in the degree to which different strains and clones underwent metacyclogenesis during growth. Examination of a reportedly avirulent L. major clone revealed that metacyclogenesis was unusually delayed and inefficient for this clone, but that those PNA- promastigotes which could be recovered from late-stationary-phase cultures were virulent for BALB/c mice. The loss of virulence associated with frequent subculture could also be attributed to a drastic diminution in metacyclogenesis potential over time. A clone which yielded over 90% PNA- promastigotes during growth within passage 1 generated fewer than 10% PNA- promastigotes during growth by passage 94. Subcloning of late-passage attenuated promastigotes yielded a clone for which no PNA- promastigotes could be generated during growth, and an infective population could not be derived from this clone. Thus, metacyclogenesis does not appear to be stable for even cloned lines of Leishmania promastigotes, and virulence comparisons between different strains and clones can be meaningfully made only if the metacyclic populations contained within the respective inocula are determined.

Complement receptor type 3 (CR3) binds to an Arg-Gly-Asp-containing region of the major surface glycoprotein, gp63, of Leishmania promastigotes.

“…7) indicate that both infective and noninfective stages use LPG and gp63 to attach to MO. This is consistent with previous observations showing that infectivity was not dependent on the ability of the parasite to bind to, and be internalized by, MO (44).…”

“…Leishmania promastigotes have been shown to differentiate in culture from a noninfective form of promastigote in actively dividing cultures, to an infective form in stationary phase cultures (43,44). This differentiation is accompanied by changes on the promastigote surface.…”

Platelet activation attracts a subpopulation of effector monocytes to sites of Leishmania major infection

“…RFP- L. major promastigotes were used for the peritoneal infection experiments. Metacyclic parasites were obtained as previously described (da Silva and Sacks, 1987) and used for all killing experiments. In brief, stationary phase parasites were washed three times with DPBS and resuspended in RPMI containing 50 µg/ml peanut agglutinin (Vector Laboratories Inc.) and incubated for 15 min at room temperature.…”

Granulocyte‐Macrophage Colony‐Stimulating Factor is a Growth‐Factor for Promastigotes of Leishmania mexicana amazonensis

“…This progressive loss of sensibility to the growth-factor upon successive passages in vitro is paralleled by a concomitant loss of infectivity and might indicate a relationship between infectivity of the promastigote and its ability to respond to the factor. A similar phenomenon occurs with cloned lines ofleishmania tropica where a decrease in the number of infective forms at each cell cycle is due to the decrease in the number of forms without receptors for peanut agglutinin [6]. The sensibility decay to the growth-factor may represent a selective advantage favouring the forms that d o not need it for their in vitro multiplication.…”