Fig 1.
Whole mounts of male (a; c; e) and female (b; d; f) Argulus japonicus showing autofluorescence in unbleached (A) and photobleached (B) specimens, and fluorescence of Phen–Green (C). (a–d) Micrographs showing the anterior region of adult male and female A. japonicus where antennules (Antl), antennae (Ant), maxillules (Ml), maxillae (Ma), and basal segment of the maxillae (Bpl) in untreated (A), photobleached (B) and Phen–Green fluorochrome (C). (e–f) Micrographs of the thoracic region of male (e) and female (f) A. japonicus. In male parasites fluorescence of untreated (A and B) and Phen–Green treated (C) reactions were present in the ventral carapace between the third and fourth swimming legs (Sl) and of the peg (P) of the podomeres of the third pair of swimming legs (Sl). In females the spines on the spermathecae (Sps) were observed for all treatments and in utero eggs were observed to produce a positive reaction for the Phen-Green (Cf). All images were taken using Zeiss bandpass filter set 01 (BP 365/12) at 490–520 nm. Scale bars represent 100 μm.
Fig 2.
Micrographs showing whole mount (A) and longitudinal sections (B, C, D and E) through male Argulus japonicus. Section planes are indicated on the light micrograph of a whole mount of a male A. japonicus (A) with dotted lines indicating sections for B–E. Fluorescence of the Phen–Green probe indicates a positive reaction for divalent cations including trace metals in longitudinal sections (B–C) of the maxillule (Ml) and maxilla (Ma). The cuticle of these regions produce more intense signals than the cuticle of the carapace (Ca) of the cephalic shield. (D) A longitudinal section through the posterior region of the thorax and abdomen shows positive fluorescence for the carapace (Ca) of the lobes of cephalic shield and the testes (Ts) in the abdomen. More intense signal was observed for the portion of the carapace of the ventral side of the thorax (Vp). (E) Longitudinal section through the thorax shows positive signals for the peg (P) of the third pair of swimming legs (Sl) and the carapace of the ventral surface of the thorax (Vp). Images were acquired using Zeiss bandpass filter set 01 (BP 365/12) at 490–520 nm.
Fig 3.
Micrographs showing whole mount (A), frontal section (B) stained with Azocarmine–Aniline blue (AZAN) and longitudinal sections treated with Phen–Green (C, D, E, F and G) through female Argulus japonicus. (A) Section planes are indicated on the light micrograph of an adult female A. japonicus with dotted lines for sections C and D; dotted line indicated labelled B indicates area where frontal section of the reproductive tract imaged in image B. (B) Frontal section of the thorax stained with AZAN containing mature oocytes (Oc) and shelled eggs (Eg); blue dotted line E corresponds with fluorescence micrograph E. (C) Section through the maxillule (Ml) of treated with Phen–Green. (D) Longitudinal section through the cephalic shield of the carapace (Ca) and antennule (Antl). (E) Longitudinal section through the thorax treated with Phen–Green showing fluorescence amorphous layer (Pm) surrounding eggs (Eg) in utero which corresponds with image G. (F) Section through the spermathaceal spine (Sps) treated with Phen–Green. (G) Section through an in utero egg (Eg) showing intense signal produced by amorphous layer (Pm) surround the egg. Images were acquired using Zeiss bandpass filter set 01 (BP 365/12) at 490–520 nm.
Fig 4.
Bar graph showing means for the natural logarithm of the weight percentages (wt%) for elements identified in sections of male (black bars) and female (grey bars) Argulus japonicus by SEM/EDS.
Error bars indicate 95% confidence intervals.