PMC9382088_01
Ultrasonographic images at the left external iliac artery (A), mid femoral artery (B), and echocardiographic images (C,D) revealed linear hyperechoic structures in the cardiovascular lumen which were subsequently molecularly confirmed to be Dirofilaria immitis.
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F1
The clinical signs, especially the lameness of both hindlimbs, persisted until the follow-up period a week later, with progressive bruise-necrosis wounds, particularly at the footpad of the right hindlimb (Supplementary Figure 1). Furthermore, hyperechoic, linear worm-like structures were found at the bifurcation of the abdominal aorta, the external iliac arteries (Figure 1A), the distal femoral arteries near the distal femur (Figure 1B), and the saphenous arteries of both hindlimbs. The 2D echocardiography images (Figure 1C) revealed an enlarged RA and RV with a flattened interventricular septum (IVS), suggesting eccentric hypertrophy and volume overload of the RV. Multiple hyperechogenic lines were observed in the RA, RV, and PA, indicating the presence of adult heartworms (Figure 1D).
The photographic images displayed the adult Dirofilaria immitis (red arrows) removal from the jugular vein (A) and the femoral artery (B). At the jugular vein, a modified, brush-like instrument (yellow arrow) was used to take off the cardiac heartworms.
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F2
Best, Netherland) using a blush-like instrument made from small diameter Kirshner wire (no. 26) with monofilament, non-absorbable polyamide suture material (size 3/0, Dafilon, B. Braun, Ruby, Spain) (Supplementary Figure 2). After twisting the instrument several times and then withdrawing it, the adult heartworms were found attached to the phlebotomy line (Figure 2A). Femoral arteries were cut linearly by a surgical blade for a 1 cm incision and alligator forceps (Microsurgical Grasping Forceps, 1.4 mm of tip width, and 6 inches of shaft length) were used to remove adult heartworms from femoral and iliac arteries (Figure 2B).
A maximum-likelihood phylogenetic tree showing the genetic relation between Dirofilaria immitis from heart, femoral artery, and wound (red color) and other filarial species using cytochrome oxidase subunit I gene.
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F3
The PCR products were subjected to gel electrophoresis, purified, and sequenced. The genetic identification revealed that all nematodes were D. immitis (Figure 3). Thickening and irregularity of the pulmonic valve and pulmonary trunk were observed (Supplementary Figure 3).
Pedal gangrenous necrosis was demonstrated in the metatarsal-phalangeal region of the hindlimb (A). Microscopically, occluding thrombus is demonstrated in the artery with the presence of a cross-section of the worm, ~100-250 mum in size (arrow) (B). Bar = 100 mum.
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F4
A significant gross lesion observed during the external examination was restricted to the paws of both hindlimbs and pedal gangrenous necrosis was demonstrated in the metatarsal-phalangeal regions (Figure 4A). Foci of infarcts were observed at the head of the spleen (Supplementary Figure 4) and the right kidney. Heartworms were also observed inside several obstructive thrombi (Figure 4B).