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Lophomonas blattarum-like organism in bronchoalveolar lavage from a pneumonia patient: current diagnostic scheme and polymerase chain reaction can lead to false-positive results

Moses Lee1orcid, Sang Mee Hwang1, Jong Sun Park2, Jae Hyeon Park3,*orcid, Jeong Su Park1,*orcid
Parasites, Hosts and Diseases 2023;61(2):202-209.
Published online: May 23, 2023

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*Correspondence: (pjh, bjack9@gmail.com; pjs, mdmicrobe@gmail.com)
• Received: September 1, 2022   • Accepted: February 16, 2023

© 2023 The Korean Society for Parasitology and Tropical Medicine

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (https://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Lophomonas blattarum-like organism in bronchoalveolar lavage from a pneumonia patient: current diagnostic scheme and polymerase chain reaction can lead to false-positive results
Parasites Hosts Dis. 2023;61(2):202-209.   Published online May 23, 2023
DOI: https://doi.org/10.3347/PHD.22107
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Lophomonas blattarum-like organism in bronchoalveolar lavage from a pneumonia patient: current diagnostic scheme and polymerase chain reaction can lead to false-positive results
Parasites Hosts Dis. 2023;61(2):202-209.   Published online May 23, 2023
DOI: https://doi.org/10.3347/PHD.22107
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Lophomonas blattarum-like organism in bronchoalveolar lavage from a pneumonia patient: current diagnostic scheme and polymerase chain reaction can lead to false-positive results
Image Image Image
Fig. 1 Light microscopy observation of wet mounted bronchoalveolar lavage fluid of the patient. (A) Multiple ciliated cells with morphological features similar to previously reported Lophomonas blattarum. (B) Normal ciliated epithelial cell with columnar shape, apical nucleus, terminal bar structure (arrow), and unidirectional insertion of cilia, from another patient’s bronchoalveolar lavage fluid sample. Scale bar=20 μm.
Fig. 2 Findings of the patient’s bronchoalveolar lavage fluid by transmission electron microscopy. (A) A dead normal bronchial ciliated epithelial cell with the nucleus in an apical position, regular unidirectional insertion of cilia, and typical terminal bar structure (white dashed box) with nearby mitochondria. (B) Dead ciliated epithelial cell with typical morphology of ciliocytopthoria. (C) Ciliated epithelial cell tightly bound to another cell with multiple mucinous vacuoles in the cytoplasm, which resembles a goblet cell in bronchial epithelium. Cellular border is shown in the magnified region (white dashed arrow). (D) Ciliated epithelial cell bound to another ciliated cell. Both cells had lost their typical morphological features. Cellular border (white dashed arrow) and tight junction (red arrow) are shown in the magnified region. N, nucleus; M, mucinous vacuoles. White arrow indicates mitochondria gathered around the ciliary insertion site. Black arrow shows cilia in a tightly bound cell. White dashed arrow demonstrates cell membrane border. Red arrow indicates tight junction. Scale bar=2 μm.
Fig. 3 Polymerase chain reaction (PCR) and Sanger sequencing results. (A) Initial gel electrophoresis results show that the previously reported Lophomonas-specific primer pair reacted to all human DNA controls and produced a 307-bp product. Negative result to Staphylococcus aureus colony DNA showed that nonspecific amplification had not occurred. (B) The results are reproducible with more human DNA. Lanes 1, 2, and 3 contained DNA extracted from bronchoalveolar lavage fluid of 3 patients without lophomoniasis. Lanes 4 and 5 contained normal DNA extracted from blood. A 251-bp product (white dashed box) appeared in addition to the 307-bp product. DW was a negative control with distilled water. (C, D) Sanger sequencing results showed 2 different amplified sequences. The 307-bp product aligned to chr11: 31907276–31907544 and the 251-bp product to chr4: 15394040–15394252 in hg19. Forward and reverse primers did not fully match the genome, but at least 11 base pairs matched the genomic sequence, including >7 consecutive matches at the 3’ end. Base of primers (highlighted) matching with the human genome are capitalized. L, Ladder; Bld, control DNA extracted from healthy blood donors. SA, PCR result for DNA extracted from a Staphylococcus aureus colony. P, DNA extracted from bronchoalveolar lavage fluid from patient suspected of having Lophomonas infection. BAL, DNA extracted from Lophomonas-negative bronchoalveolar lavage fluid from another patient. DW, distilled water.

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Lophomonas blattarum-like organism in bronchoalveolar lavage from a pneumonia patient: current diagnostic scheme and polymerase chain reaction can lead to false-positive results
PHD : Parasites, Hosts and Diseases

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