Fig 1.
The study experimental design.
Fig 2.
TEM micrographs of the prepared nano-formulations showing spherical shape with discrete separation.
(A) Blank NLCs (scale bar = 500nm). (B) NTZ loaded NLCs (scale bar = 200nm).
Fig 3.
In vitro NTZ release profile from NTZ-loaded NLCs at 37 °C.
Table 1.
Oocyst count and parasite percent reduction among infected immunocompetent subgroups at different examination times.
Table 2.
Oocyst count and parasite percent reduction among infected immunosuppressed subgroups at different examination times.
Table 3.
Comparison between the immunocompetent and immunosuppressed groups regarding oocyst count at different examination times.
Fig 4.
Oocysts count and parasite percent reduction in stool samples among different infected immunocompetent (A and C) and immunosuppressed (B and D) subgroups.
Fig 5.
(A) and (B) Modified Ziehl Neelsen-stained stool samples revealing Cyclospora oocysts (X 400).
Fig 6.
(A) and (B) Safranin-stained stool samples revealing Cyclospora oocysts (X 400).
Fig 7.
SEM of Cyclospora oocysts collected from different studied subgroups of the immunocompetent murine model.
(A) Oocyst from subgroup I b (infected untreated control), revealing spherical shape with smooth regular surface (X 22,000). (B) Oocyst from subgroup I c (receiving blank NLCs), showing spherical smooth surface organism (X 22,000). (C) Oocyst from subgroup I d (treated with TMP-SMX), showing cauliflower-like shrunken parasite losing its smooth surface with noticeable furrows and ridges (X 22,000). (D) Oocyst from subgroup I e (treated with NTZ), showing a swollen distorted parasite losing the smooth surface with apparent surface erosion and ulceration (X 8,000). (E) and (F) Oocysts from subgroup I f (treated with NTZ-loaded NLCs), showing mutilated rupturing parasite losing its spherical shape and smooth surface with multiple blebs, clefts, lacerations and profound cavitations on the parasite surface (X 8,000 and 5,000).
Fig 8.
SEM of Cyclospora oocysts collected from different studied subgroups of the immunosuppressed murine model.
(A) Oocyst from subgroup II b (infected untreated control), showing a spherical-shaped organism with smooth regular surface (X 22,000). (B) Oocyst from subgroup II c (receiving blank NLCs), demonstrating spherical smooth surface parasite (X 22,000). (C) Oocyst from subgroup II d (treated with TMP-SMX), revealing dimpled irregular outer surface (X 22,000). (D) Oocyst from subgroup II e (treated with NTZ), showing a disorganized organism with noticeable multiple deep perforations (X 22,000). (E) and (F) Oocysts from subgroup II f (treated with NTZ-loaded NLCs), showing a disfigured rupturing parasite losing its spherical shape and smooth surface with deep cavitations (X 8,000 and 5,000).
Table 4.
Comparison between the different studied immunocompetent subgroups according to intestinal villi length on the 14th and 30th days P.I.
Fig 9.
Intestinal histopathologic changes of immunocompetent group on the 14th and 30th days P.I. (H&E, x100, scale bar = 200 μm).
14th day P.I: (A) Normal intestinal histology is seen in normal uninfected control group. (B) Infected untreated subgroup showing short (bi-headed arrow) broad edematous villi with dilated capillaries (arrow head) as well as severe inflammation (*). Inset (x1000) shows the organism within enterocyte (red arrow). (C) The subgroup receiving blank NLCs demonstrating similar histopathological findings as in infected untreated control. (D) TMP-SMX treated subgroup reveals increased villous length (bi-headed arrow) with improved inflammation. (E) In NTZ treated subgroup, increased villous length (bi-headed arrow) is observed but moderate inflammation is still present (*) as well as congested capillaries (arrow head). (F) NTZ-loaded NLCs subgroup presents long slender villi (bi-headed arrow) with no significant inflammation. 30th day post infection: (G) Normal histology is apparent in normal uninfected subgroup. (H) and (I) The pathologic changes are exaggerated in infected untreated control and blank NLCs subgroups respectively. (J) In TMP-SMX subgroup, moderate inflammation is noticed back again (*). (K) In NTZ treated subgroup, moderate inflammation is seen (*). (L) NTZ-loaded NLCs subgroup shows restoration of normal villous length and lack of inflammation. Bi-headed arrow = villous length, * = inflammation, arrow head = dilated capillaries.
Table 5.
Comparison between the different studied immunosuppressed subgroups according to the intestinal villi length on the 14th and 30th days P.I.
Fig 10.
Intestinal histopathologic changes of immunosuppressed group on 14th and 30th days P.I. (H&E, x100, scale bar = 200 μm).
In 14th days post infection: (A) Normal uninfected control reveals normal intestinal architecture. (B) and (C) Both infected untreated control and blank NLCs subgroups present short (bi-headed arrow) broad edematous villi as well as inflammatory changes (*). (D) TMP-SMX treated mice shows increased villous length with moderate inflammation in some villi (*). (E) NTZ treated subgroup displays increased villous length, however moderate inflammation is still existing (*). (F) The subgroup treated with NTZ-loaded NLCs demonstrates restoration of the normal long slender villous morphology. In 30th days post infection: (G) Normal histology is still seen in normal uninfected group. (H) and (I) More apparent pathologic changes of Cyclospora infection are noticed in infected untreated and blank NLCs subgroups respectively. (J) Residual inflammation (*) is still observed in some villi in TMP-SMX treated mice. (K) NTZ treated subgroup shows similar pathologic findings as TMP-SMX. (L) Restoration of villous length is still obviously noted in NTZ-loaded NLCs with no evident inflammation. Bi-headed arrow = villous length, * = inflammation.