Molecular characterization of Trichomonas tenax causing pulmonary infection.

A 58-year-old man presented with retrosternal pain 30 months after a surgical operation for reflux esophagitis in relation to Barrett’s esophagus. Gastroscopy revealed an ulcer of the inferior part of the esophagus, which on biopsy proved to be an adenocarcinoma. Additional surgery was performed through the abdomen and through the thorax via the sixth intercostal space. Resection of the lower part of the esophagus and high anastomosis were performed because of nondilatable peptic stenosis. Two chest tubes were put in place: one draining the right pleura and the other in contact with the anastomosis. On the second postoperative day, a sepsis-like syndrome including bronchospasm developed with increasing ventilation requirements. A chest radiograph revealed no abnormalities, and a perfusion computed tomography (CT) scan did not show pulmonary thromboembolism. Amoxicillin-clavulanic acid and gentamicin were given for 8 days despite various bacteriological analyses that were all sterile, including 10 blood samples, 3 pleural fluid samples, 1 bronchoalveolar lavage, 1 bronchial aspiration, and 1 urine sample. The patient improved partially, and the pleural tube was removed. On the 13th postoperative day, the patient was still ventilator dependent. Fever recurred, and biological investigations showed a leukocyte count of 11,700/mm and a C-reactive protein level of 128 mg/liter. A chest CT scan revealed a small collection of 1 cm in diameter in contact with the posterior part of the anastomosis and a new infiltrate of the left lung parenchyma located in the apical segment of the lower lobe. Analysis of the fluids from both bronchoalveolar lavage and chest tube showed a large number of neutrophils and numerous flagellated, motile organisms with the typical appearance of trichomonads, visible on wet preparations and Giemsa staining. Culture of bronchoalveolar fluid yielded Neisseria lactamica, while culture of tube fluid yielded alphaand beta-hemolytic streptococci and Haemophilus parainfluenzae. Treatment with metronidazole, piperacillintazobactam, and gentamicin was started. Trichomonads were not found in the drained fluid, and a chest CT scan showed the resolution of the collection and no change in pulmonary parenchyma 3 and 5 days after the initiation of metronidazole therapy, respectively. The CT scan failed to show passage of contrast material from the esophageal suture line into the mediastinum. Antibiotics, including metronidazole, were continued for 10 days, and the tube was removed later. However, the patient had remained ventilator dependent since the operation. Ischemic brain damage of unknown origin was revealed by cerebral magnetic resonance imaging, and the patient remained in a deep coma for 1 week and then died. In order to identify the species of Trichomonas, we amplified directly from the bronchoalveolar fluid and sequenced the 5.8S rRNA gene and the internal transcribed spacer flanking regions (ITS1 and ITS2) with the forward and reverse primers TRICHO-F and TRICHO-R (5 -CGGTAGGTGAACCTGC CGTT-3 and 5 -TGCTTCAGTTCAGCGGGTCT-3 , respectively) as described previously (6). DNA extraction from clinical specimens and PCR were performed in a protected molecular biology area. Negative (bronchoalveolar fluids) and positive (vaginal specimen with Trichomonas vaginalis) controls were included in the series. Sequence analysis of both strands was carried out on a 3700 DNA analyzer (Applied Biosystems, Courtaboeuf, France). The sequence of the amplified 368-bp fragment was compared with the National Center for Biotechnology Information database sequences and exhibited 100% homology to Trichomonas tenax (accession no. U86615). The sequence had only 96% identity to Trichomonas gallinae (16 nucleotide differences in the 368-bp fragment), 93% identity to T. vaginalis (25 nucleotide differences in the 368-bp fragment), and 85% identity to Pentatrichomonas hominis (51 differences in the 339-bp fragment).