Ciliocytophthoria in Clinical Virology.

Direct immunofluorescence assays (DFAs) are commonly used in the clinical virology laboratory for the rapid detection of a wide variety of viral pathogens. These rapid and highly specific assays consist essentially of staining an air-dried portion of a patient specimen with a fluorescein-labeled antibody that is directed toward a specific viral pathogen. After removing unbound stain by washing, the specimen is examined using fluorescence microscopy for the presence of virally infected cells. To ensure that adequate and appropriate cells are present for staining, the cellularity of specimens submitted for DFA should be assessed. During this assessment, clinical virologists may encounter unfamiliar cellular elements or cellular fragments.

Ciliocytophthoria have been misinterpreted as ciliated Microorganisms.[1-4] Hilding first noted these anucleate, apical remnants of ciliated epithelial cells in 1930.[5] In 1956, Papanicolaou used the term ciliocytophthoria to describe these degenerative cellular fragments.[6] Ciliocytophthoria are now well described by cytologists and cytopathologists and are most often seen in fixed, stained cytologic preparations.[1,3-11] Motile forms have also been observed in flesh nasopharyngeal and peritoneal specimens.[2,4]

We describe both motile and nonmotile ciliocytophthoria in a fresh specimen, which originally raised concerns of a parasitic infection. We discuss means to differentiate this entity from the pathogenic-ciliated protozoa Balantidium coli. It is important that clinical virologists, who now examine the cytologic contents of some specimens, are familiar with ciliocytophthoria and are able to avoid this potential diagnostic pitfall.

REPORT OF A CASE

A 7-month-old male infant, who was born premature at 30 weeks' gestation, presented to a local pediatrician with wheezing. A viral origin was suspected, and a nasopharyngeal swab was submitted in M4 media to the microbiology laboratory. A DFA for respiratory syncytial virus was requested.

The medical technologist prepared a wet mount of the specimen to determine if an adequate number of columnar epithelial cells were present. The examination of the wet mount, however, revealed a round-to-oval, rapidly moving, but dyssynchronous, ciliated form that was thought to represent a ciliated microorganism (Figure 1). Numerous morphologically similar but nonmotile forms were also noted. The specimen was then examined by parasitologists and pathologists, who determined the forms to represent ciliocytophthoria and not a ciliated protozoan. A Diff-Quik stain confirmed the identification of ciliocytophthoria (Figure 2). Both respiratory syncytial virus DFA and additional viral cultures for other respiratory pathogens were negative. The patient was seen in a follow-up visit 5 days later, and he appeared healthy.

[Figure 1-2 ILLUSTRATION OMITTED]

COMMENT

The enumeration and differentiation of cells present in clinical specimens submitted for microbiologic studies have already proven cost-effective and medically relevant.[12] For instance, it is now standard of practice to determine the number of squamous epithelial cells in specimens submitted for bacterial sputum culture and to reject specimens contaminated with oropharyngeal flora.[13] In clinical virology, it is important to assess a specimen submitted for DFA to determine the presence and number of columnar epithelial cells. Specimens with few columnar epithelial cells may be suboptimal for DFA, since potentially infected cells either are not present or may be present in low numbers.[14] In our laboratory, if fewer than 25 ciliated respiratory epithelial cells are present, the specimen is processed, but the clinician is notified that the specimen may be insufficient for the detection of virus by DFA.

During the examination of a respiratory specimen for cellular constituents, virologists may encounter unfamiliar cells, cell fragments, Curschmann spirals, and artifacts, some of which simulate microorganisms. Probably the most confusing are ciliocytophthoria, the ciliated, apical anucleate fragments of respiratory epithelial cells. These cellular fragments are often round, smaller than intact epithelial cells, and usually encountered in specimens taken from anatomic locales that are lined by ciliated columnar epithelium. Ciliocytophthoria have been reported in respiratory tract specimens, gynecologic specimens, and peritoneal washings.[1-3,5,6,8-11,15] Ciliocytopthoria have been produced experimentally by exposing porcine respiratory epithelium to a wide variety of pathogens and are known to be associated with respiratory disease in horses.[16,17] In humans, ciliocytophthoria have been associated with acute tonsillitis and viral infections.[18-20] Clinical virologists should, therefore, expect these pseudoparasites in respiratory specimens submitted for viral DFA.

In a wet preparation the presence of ciliocytophthoria can be disconcerting to those unfamiliar with this entity, especially if beating cilia are seen. In fresh specimens, cellular tufts may retain cilial motility for several hours.[21] There are, however, several clues that these, often motile, cellular fragments represent ciliocytophthoria rather than a ciliated parasite. The microcopist examining the specimen should consider the size of the suspected "organism" and the distribution of the cilia and should determine if a nucleus is present. Using these criteria, ciliocytophthoria may be successfully differentiated from the only human parasitic ciliate, B coli. High-magnification microscopy is usually sufficient to confirm ciliocytophthoria (Figure 3). If doubt still remains, special stains may be used to confirm the anucleate nature of these structures and to accentuate the cilia (Figure 2).

[Figure 3 ILLUSTRATION OMITTED]

Infections due to B coli are extremely rare and usually involve the large intestine rather than the respiratory tract.[22] Balantidium coli are the largest single-celled human parasite and measure 50 to 100 x 40 to 70 [micro]m.[22] Ciliocytophthoria are much smaller; in our specimen, they averaged 12 x 10 [micro]m. Ciliocytophthoria usually demonstrate cilia predominantly along one edge, but if viewed on end they may appear to be circumferentially ciliated.[1] Balantidium coli are uniformly covered with cilia.[21,22] Although dyssynchronous cilial motility was seen in this case, ciliocyiophthoria have also been noted to produce rhythmic motility.[23] Finally, ciliocytophthoria are anucleate (Figures 1 through 3), whereas B coli have a macronucleus that is typically indented or kidney shaped (Figure 4).[21,22] A careful inspection for these features should allow for accurate morphologic differentiation between ciliocytophthoria and B coli. Ashfaq-Drewett et al found size and internal structures to be the most useful features for differentiating B coli from ciliocythophthoria.[4]

[Figure 4 ILLUSTRATION OMITTED]

Sidaway et al found ciliocytophthoria difficult to detect in air-dried, Diff-Quik-stained smears.[23] We, however, found this rapid stain easy to perform an interpret and useful for the confirmation of ciliocytophthoria on an air-dried aliquot of specimen. In the Diff-Quick-stained smear, it became apparent that the cellular fragments were indeed anucleate when compared with nearby nucleated respiratory epithelial cells (Figure 2). Furthermore, the air-dried cell fragments lay flatter in the stained smear and more obviously demonstrated cilia associated with only one edge. Finally, the Diff-Quik method demonstrates the terminal bar (Figure 2), a feature typical of ciliated respiratory epithelium.

As clinical virologists more frequently examine the cellularity of respiratory specimens, they should expect to encounter these and other cellular fragments. Ciliocytophthoria should be considered when round-to-oval, motile, or nonmotile ciliate forms are identified in clinical specimens from anatomic sites that are lined by ciliated epithelial cells. The absence of a nucleus and the presence of cilia predominantly along one edge is characteristic. The Diff-Quik stain is a rapid method that may be used to confirm the distribution of the cilia and the anucleate character of these cellular fragments. It is also an excellent method for demonstrating the terminal bar of ciliated respiratory epithelium. When doubt remains, an experienced cytologist, pathologist, or parasitologist should review the smear.

References

[1.] Mahoney CA, Sherwood N, Yap EH, Singleton TP, Whitney DJ, Cornbleet PJ. Ciliated cell remnants in peritoneal dialysis fluid. Arch Pathol tab Med. 1993; 117:211-213.

[2.] Nosanchuk JS. Pseudoparasites due to Ciliocytophthoria. Arch Pathol Lab Med. 1993;117:967.

[3.] Roxby CM, Wood M, Martin AM, McHugh M. 1993. Ciliated organisms seen in fluid following dialysis. Lancet. 1993;1:916.

[4.] Ashfaq-Drewett R, Allen C, Harrison RL. Detached ciliary tufts: comparison with intestinal protozoa and a review of the literature. Am J Clin Pathol. 1989; 93:541-545.

[5.] Hilding A. The common cold. Arch Otolaryngol. 1930;12:133-150.

[6.] Papanicolaou GN. Degenerative changes in ciliated cells exfoliating from the bronchial epithelium as a cytologic criterion in the diagnosis of diseases of the lung. N Y J Med. 1956;56:2657.

[7.] Johnston WW, Elson CE. Respiratory tract. In: M. Bibbo, ed. Comprehensive Cytopathology. Philadelphia, Pa: WB Saunders Co; 1991:340-341.

[8.] Hollander DH, Gupta PK. Detached ciliary tufts in cervico-vaginal smears. Acta Cytol. 1974;18:367-369.

[9.] Kanbour A, Doshi N. Psammoma bodies and detached ciliary tufts in a cervicovaginal smear associated with benign ovarian cystadenofibroma. Acta Cytol. 1980;24:549-552.

[10.] Clocuh YP. Ciliocytophthoria in pulmonary and vaginal cytology [in German]. Medizinische Welt. 1978;29:1044-1046.

[11.] Clocuh YP. Ciliocytophthoria in the cervical smear [in German]. Geburtshilfe Frauenheilkd. 1978;38:229-230.

[12.] Washington JA. The clinical microbiology laboratory: utilization and cost-effectiveness. Am J Med. 1985;78:8-16.

[13.] Murray PR, Washington JA. Microscopic and bacteriologic analysis of expectorated sputum. Mayo Clin Proc. 1975;50:339-344.

[14.] Lyerla HC. Diagnostic applications of immunofluorescence tests in the virology laboratory, in: Lenette DA, Specter S, Thompson K, eds. Diagnosis of Viral Infections. Baltimore, Md: University Park Press; 1979:103-113.

[15.] Hubel E, Kanitz M, Kuhlmann U. Ciliocytophthoria in peritoneal dialysis effluent. Peril Dial Int. 1990; 10:179-180.

[16.] Williams PP, Gallagher JF, Pirtle EC. Effects of microbial isolates on porcine tracheal and bronchial explant cultures as observed by scanning electron microscopy. Scan Electron Microsc. 1981;4:141-150.

[17.] Freeman KP, Roszel JF, Slusher SH. Inclusions in equine cytologic specimens. J Am Vet Mad Assoc. 1985;186:359-364.

[18.] Sasaki Y, Abe H, Tokunaga E, Tsuzuki T, Fujioka T. Ciliocytophthoria (CCP) in nasopharyngeal smear from patients with acute tonsillitis. Acta Oto. 1988; 454(suppl):175-177.

[19.] Sasaki Y, Korematsu M, Naganuma M. Ciliocytophthoria (CCP)in nasal secretions: relation of viral infection to otorhinological disease [in Japanese]. Josai Shika Daigaku Kiyo. 1987;16:441-445.

[20.] Grabowska-Joachimiak A. The morphology of ciliated cells in nasal mucosa during a viral infection [in Polish]. Otolaryngol Pol. 1998;52:175-180.

[21.] Ash L, Orihel T. Atlas of Human Parasitology. 4th ed. Chicago, Ill: ASCP Press; 1997:116-117, 370-371.

[22.] Garcia LS, Bruckner DA. Intestinal protozoa: flagellates and ciliates. In: Diagnostic Medical Parasitology. 3rd ed. Washington, DC: ASM Press; 1997:49-51.

[23.] Sidaway MK, Poonam C, Oertel YC. Detached ciliary tufts in female peritoneal washings: a common finding. Acta Cytol. 1987;31:841-844.

Accepted for publication December 6, 1999.

From the Section of Clinical Microbiology, Department of Clinical Pathology, Division of Pathology and Laboratory Medicine, Cleveland Clinic Foundation, Cleveland, Ohio.

Reprints: Gary W. Procop, MD, Clinical Microbiology/L40, The Cleveland Clinic Foundation, 9500 Euclid Ave, Cleveland, OH 44195.