Microbiology Case Study: A 57 Year Old Man with Fever

Case History

A 57 year old male with a recent history of a left above the knee amputation developed a fever during the same admission of 101.1°F. His amputation had been complicated by poor wound healing, and he had a simultaneous right leg abscess that grew methicillin-sensitive Staphylococcus aureus. Examination of his wound showed serosanguinous drainage with no erythema or purulence. Blood cultures and a wound swab were sent for microbiological analysis.

Laboratory Findings Wound cultures grew methicillin-resistant Staphyloccocus aureus thought to represent colonization rather than true infection. Blood cultures flagged positive in one anaerobic bottle only at 30 hours. A gram smear showed gram-negative cocci (Image 1). Anaerobic blood plates grew pinpoint colonies (Image 2). MALDI-TOF identified the bacteria as a Veillonella species.

Image 1. Gram stain from anaerobic culture showing gram negative cocci.
Image 2. Growth on anaerobic blood plate.

Discussion

Veillonella species are gram negative cocci. They are lactate fermenting, obligate anaerobes and are considered normal flora of the intestines and oral mucosa. As such, they are usually regarded as a contaminant. They have, however, been implicated in osteomyelitis, prosthesis infections, and endocarditis. They are particularly associated with poor oral hygiene, chronic periodontitis, and smoking. They have important implications in dental disease due to their ability to form biofilms. They are frequently resistant to ampicillin and have also been noted to be resistant to tetracyclines in periodontal patients. Identification is done by molecular methods, typically MALDI-TOF. PCR has also been developed, but is not routinely used.

This was considered a contamination due the absence of symptoms and isolation in one bottle only. A follow up blood culture was negative. Routine wound care was resumed.

References

  1. Rovery C, Etienne A, Foucault C, Berger P, Brouqui P. Veillonella montpellierensis endocarditis. Emerg Infect Dis. 2005;11(7):1112–1114.
  2. Mashima I, Theodorea CF, Thaweboon B, Thaweboon S, Nakazawa F. Identification of Veillonella Species in the Tongue Biofilm by Using a Novel One-Step Polymerase Chain Reaction Method. PLoS One. 2016;11(6):e0157516. Published 2016 Jun 21.

-Jonathan Wilcock, MD is a 1st year anatomic and clinical pathology resident at the University of Vermont Medical Center.

-Christi Wojewoda, MD, is the Director of Clinical Microbiology at the University of Vermont Medical Center and an Associate Professor at the University of Vermont.

Surgical Pathology Case Study: A 6 Year Old Patient with Sudden Onset Abdominal Pain and a Worrisome Mass on Imaging

Case History

The patient is a 6 year old who developed abdominal pain 2 days prior to admission. The patient was in school when the abdominal pain began, resulting in the patient doubling over in pain. The pain resolved within 1 hour, however, because the initial presentation was an unremitting abdominal pain, the patient was taken to an outside hospital for evaluation. There was no vomiting, diarrhea, or constipation. On physical exam, the patient was very tender to palpation in the right lower quadrant and was unable to tolerate deep palpation. A computed tomography scan was subsequently ordered which showed a large mass in the pelvic peritoneum. The patient was admitted to surgery for an exploratory laparotomy, with resection of the pelvic mass.

Diagnosis

Received fresh in the Surgical Pathology laboratory is a 162.5 gm, 10.2 x 7.5 x 4.0 cm lobulated, ovoid mass of pink-tan, rubbery tissue that appears encapsulated by a thin translucent membrane. The margins are inked black and the specimen is serially sectioned revealing glistening, gray-tan soft tissue with focal areas of yellow discoloration and softening. Along one edge of the specimen, there is a 4.0 x 1.5 cm rim of dark red-brown, rubbery tissue (Figure 1). Portions of the fresh specimen are submitted in glutaraldehyde for electron microscopy if needed, RPMI for cytogenetics, and are snap-frozen as well. Touch preparations are also made and gross photographs are taken. Representative sections are submitted as follows:

Cassette 1-7:    Sections of mass including inked capsule

Cassette 8-10:   Representative sections from central portion of mass including areas of softening and discoloration

Cassette 11-13: Additional representative sections of the mass

Image 1. Cut surface of a gray-tan mass with yellow areas of discoloration and hemorrhage around periphery.

Histologically, the mass is composed of sheets and nests of small round cells along thin fibrous septa, giant multinucleated cells, and rare strap cells. Necrosis less than 5%. The margins are positive, although the specimen is unoriented. Venous and lymphatic invasion is absent. Immunohistochemical (IHC) stains are ordered and the results are listed below:

Positive IHC stains: Myogenin, desmin, CD56 and Bcl-2

Negative IHC stains: S-100, keratin AE1/AE3, CAM 5.2, SMA, CD99, Fli-1, WT-1, and EMA

In addition to the IHC stains, a portion of tissue was sent for cytogenetics testing, which showed a chromosomal translocation at t(2;13)(q35;q14). Based on the histologic appearance, IHC stains, and cytogenetic testing, the specimen was signed out as an alveolar rhabdomyosarcoma with a pathologic stageof pT2b, N0, MX.

Following the diagnosis, the patient was placed on a chemotherapy regimen of Vincristine, Adriamycin, Etoposide and Cytoxan, as well as radiation therapy.

Discussion

Rhabdomyosarcoma is the most common malignant soft tissue tumor in children and is the most common malignant solid tumor in children after neuroblastoma and Wilms tumor, accounting for 5-10% of all childhood tumors. 90% of these tumors occur in patients under the age of 25, and approximately 70% occur in children under 10 years of age. The most common locations of rhabdomyosarcoma are in the head and neck region, followed by the genitourinary system, extremities and then torso.

The 2013 World Health Organization classification of skeletal muscle tumors divided rhabdomyosarcoma into four types based on histology:

  1. Embryonal rhabdomyosarcoma (botryoides and anaplastic variant)
  2. Alveolar rhabdomyosarcoma (solid and anaplastic variant)
  3. Pleomorphic rhabdomyosarcoma
  4. Spindle cell/sclerosing rhabdomyosarcoma

Alveolar rhabdomyosarcoma (ARMS) accounts for approximately 20-30% of all rhabdomyosarcoma tumors, with no genetic predisposition. Although it is most common in teenagers, ARMS affects all ages. Most patients will present with a painless soft tissue mass, but based on the size and location of the mass, it may cause mass effect. A quarter of patients will have metastasis at the time of diagnosis, most commonly to the bone marrow, bones, and lymph nodes.

Grossly, ARMS presents as a solid, well-defined mass with a fleshy, tan-gray cut surface. Histologically, it is composed of small, blue, round cells and occasional round to spindle shaped rhabdomyoblasts. When compared to embryonal rhabdomyosarcoma, the rhabdomyoblasts in ARMS are slightly larger. ARMS is broken down into two subtypes: the classic subtype and the solid subtype. In the classic subtype, the tumor is composed of nests of cells that adhere to the edges of fibrous septa, resembling pulmonary alveoli (hence the name alveolar rhabdomyosarcoma). Multinucleated giant cells with a peripherally located nuclei may also be present. In the solid subtype, there will be nests and sheets of neoplastic cells that are separated by thin fibrovascular septa, but will not form in the classic alveolar pattern (Image 2).

Image 2. 20x photomicrograph demonstrating the neoplastic cells lining up along thin fibrous septa, giving the appearance of pulmonary alveoli

Due to the various appearances of rhabdomyosarcoma, it has become important to integrate immunohistochemical (IHC) stains and molecular testing into the diagnosis. The most common IHC stains that are used to determine the rhabdomyoblastic differentiation of a sarcoma is through the use of Myogenin and Myogenic differentiation 1 (MyoD1) stains, in which both stains will be positive in rhabdomyosarcoma. These two stains can be furthered used to help narrow down a diagnosis of ARMS because if more than 50% of the neoplastic cells express Myogenin, this is highly suggestive of a diagnosis of ARMS (Figure 3). In ARMS, the MyoD1 will have a variable expression. Additional positive IHC stains for ARMS can include: desmin, P-cadherin, and bcl-2.

Image 3. Myogenin IHC stain demonstrating a strong, homogenous expression

To go along with IHC stains, molecular testing has been shown to be affective with determining the type of rhabdomyosarcoma. There have been two translocations that have been identified in ARMS. The first is at t(2;13)(q35;q14), which results in a fusion of the PAX3 gene with the FOXO1 gene (previously known as the FKHR gene). This translocation is present in 60% of all ARMS cases, and has been found to occur mostly in older children and younger adults. The second translocation is at t(1;13)(p36;q14), which results in a fusion of the PAX7 gene with FOXO1, and is present in approximately 20% of all ARMS cases. The remaining 20% are fusion negative, and are associated with the solid subtype histologically. There is early preliminary data that shows a less aggressive disease course in patients with the PAX7-FOXO1 fusion, compared to those with the PAX3-FOXO1 fusion.

In order to determine the best treatment course, patients who are diagnosed with rhabdomyosarcoma are divided into a low risk, intermediate risk or high risk group based on the pathologic stage, clinical stage and clinical group. The pathologic stage is determined using the Pretreatment TNM Staging System that was set forth by the Intergroup Rhabdomyosarcoma Study (IRS) group (not the same as the TNM staging system put out by the American Joint Committee on Cancer) below:

The clinical stage is then determined using the TNM staging above and the Pretreatment Clinical Staging System below that is also put out by the IRS group:

In the above Clinical Staging System, a favorable site is defined as occurring in the orbit, biliary tract, head and neck region (excluding parameningeal) and genitourinary region (excluding prostate and bladder). Any other site not listed is considered unfavorable. Next, a clinical group is assigned based on the extent of the disease using the Clinical Grouping System below, which again is put out by the IRS group:

Lastly, based on the clinical stage and clinical group determined above, the patient is assigned a risk group of either low risk, intermediate risk, or high risk using the Children’s Oncology Group guidelines listed below:

When compared to embryonal rhabdomyosarcoma, which is the most common type of rhabdomyosarcoma, ARMS has a worst prognosis. The IRS group clinical group and stage can help to predict the overall outcome of the patient, with the standard treatment regimen composed of surgery, radiation therapy and chemotherapy.

References

  1. Dziuba I, Kurzawa P, Dopierala M, Larque A, Januszkiewicz-Lewandowska D. Rhabdomyosarcoma in Children – Current Pathologic and Molecular Classification. Pol J Pathol. 2018;69(1):20-32. doi:10.5114/pjp.2018.75333
  2. Liu H, Zhao W, Huang M, Zhou X, Gong Y, Lu Y. Alveolar rhabdomyosarcoma of nasopharynx and paranasal sinuses with metastasis to breast in a middle-aged woman: a case report and literature review. Int J Clin Exp Pathol. 2015;8(11):15316–15321. Published 2015 Nov 1.
  3. Owosho AA B Ch D, Huang SC Md, Chen S Mbbs, et al. A clinicopathologic study of head and neck rhabdomyosarcomas showing FOXO1 fusion-positive alveolar and MYOD1-mutant sclerosing are associated with unfavorable outcome. Oral Oncol. 2016;61:89–97. doi:10.1016/j.oraloncology.2016.08.017
  4. Ozer E. Alveolar Rhabdomyosarcoma. Pathology Outlines. http://www.pathologyoutlines.com/topic/softtissuealvrhabdo.html. Revised March 26, 2019. Accessed July 26, 2019.
  5. Rudzinski ER, Anderson JR, Hawkins DS, Skapek SX, Parham DM, Teot LA. The World Health Organization Classification of Skeletal Muscle Tumors in Pediatric Rhabdomyosarcoma: A Report From the Children’s Oncology Group. Arch Pathol Lab Med. 2015;139(10):1281–1287. doi:10.5858/arpa.2014-0475-OA
  6. Rhabdomyosarcoma Staging and Clinical Risk Groups. Stanford Medicine Surgical Pathology Criteria. http://surgpathcriteria.stanford.edu/srbc/rhabdomyosarcoma/staging.html. Accessed August 10, 2019

-Cory Nash is a board certified Pathologists’ Assistant, specializing in surgical and gross pathology. He currently works as a Pathologists’ Assistant at the University of Chicago Medical Center. His job involves the macroscopic examination, dissection and tissue submission of surgical specimens, ranging from biopsies to multi-organ resections. Cory has a special interest in head and neck pathology, as well as bone and soft tissue pathology. Cory can be followed on twitter at @iplaywithorgans.

Microbiology Case Study: An 83 Year Old Male with Fever

Case History

The infectious disease service was consulted on an 83 year old male for fever.

His past medical history was significant for diabetes mellitus, anemia and renal insufficiency. He initially presented 3 weeks ago with chills, rigors and fever to 103 degrees Fahrenheit. For the past several months, the patient has had weight loss (10-20 pounds over an unspecified timeframe), fatigue and new iron deficiency anemia. A heart murmur was heard on physical exam. The patient was admitted for suspicion of sepsis and he was started on empiric antibiotics vancomycin and ceftriaxone. Three sets of blood cultures were drawn prior to initiation of antibiotics, which were all positive for gram positive cocci in pairs and chains. Transesophageal echocardiogram (TEE). TEE showed large vegetation on posterior mitral leaflet measuring 1cm x 1.8 cm, and a smaller mass on the anterior leaflet. A week after admission, a mitral valve replacement was performed followed and a portion of the valve was sent for culture (Figure 1).

Laboratory Identification

Image 1. Gram stain from mitral valve specimen showing a large accumulation of gram positive cocci in chains (100x magnification).
Image 2. Brown-Hopps stain of the surgically resected mitral valve tissue vegetation showing sheets of gram positive bacteria (40x magnification).

Discussion

The gram positive organism from blood and mitral valve culture was identified as Streptococcus mitis by MALDI-TOF mass spectrometry. S. mitis is a member of the Streptococcus genus. Streptococci have a number of features that aid in laboratory identification: they are Gram positive, catalase-negative, spherical/ovoid, with organisms that are usually found in chains. They are facultative anaerobes.

More specifically, S. mitis belongs to the viridans streptococci group which includes Streptococcus mutans, Streptococcus sanguis, and Streptococcus salivarius, among many others. The most common infection caused by viridans streptococci is bacterial endocarditis, as in the case of this patient. Other infections can include brain abscesses, liver abscesses, dental caries, and bacteremia.

Patients with bacterial endocarditis have an infection of the heart valves or the endoecardial wall that leads to formations of vegetations. These vegetations are composed of thrombotic debris and organisms (Image 2), often associated with destruction of cardiac tissue. Its onset often involves severe symptoms including fever, chills, and weakness. Fever is the most consistent symptom of infective endocarditis, but it may be subtle or even absent in some cases, especially in older adults. Weight loss and flu-like symptoms may also be seen. Left-sided infective endocarditis, as in the case of our patient, will present with murmur in 90% of cases. In long-standing infective endocarditis, patients may present with Roth spots (retinal hemorrhages), Osler nodes (subcutaneous nodules in the digits), microthromboemboli (which appear as splinter hemorrhages under fingernails and toenails), and Janeway lesions (red nontender lesions on the palms or soles).

In the laboratory, the diagnosis of S. mitis and other viridians streptococci is often detected via blood culture as in the case of this patient. Once the blood culture bottle becomes positive, a Gram stain is performed, which shows Gram positive cocci in chains (Image 1). These features are helpful in differentiating Streptococcus from Staphylococcus (which appears as clusters instead of chains). Biochemical testing can be done to narrow down the species and identify S. mitis, which is optochin resistant (as opposed to S. pneumonia), acetoin negative (in contrast to most other viridans organisms), and urease negative (which differentiates it from S. vestibularis which is urease positive).

Surgical pathology can also aid in diagnosis by microscopically identifying vegetations on the affected valve (Image 2). Treatment of bacterial endocarditis is usually with penicillin or ceftriaxone, however susceptibility testing should be performed on S. mitis and other viridians streptococci because resistance can occur to penicillin. Blood cultures are followed until they are negative for 72 hours. In the case of our patient, his cultures became negative shortly after he started treatment. Susceptibility testing showed that the organism is sensitive to penicillin and ceftriaxone. The patient was continued on ceftriaxone and is clinically improving.

-Haytham Hasan, MD, is an Anatomic and Clinical Pathology resident at NorthShore Evanston Hospital (University of Chicago).

-Erin McElvania, PhD, D(ABMM), is the Director of Clinical Microbiology NorthShore University Health System in Evanston, Illinois. Follow Dr. McElvania on twitter @E-McElvania. 

Microbiology Case Study: An 8 Year Old Male with Left Knee Pain

Case History

An 8 year old male with no significant medical history presented with left knee pain and swelling for one week. Physical examination revealed a temperature of 101.2°F and a left swollen, tender knee with reduced range of motion. A joint aspirate was performed, and synovial fluid and blood were sent for microbiological analysis.

Laboratory Findings

Synovial fluid analysis showed increased neutrophils, a nucleated cell count of 90,840 cells/cmm, and no crystals.

Blood cultures were negative. Gram smear of the joint fluid showed many neutrophils and no bacteria. Fluid culture grew convex tan-yellow colonies on blood and chocolate plates at 48 hours (Image 1). Gram smear revealed gram-negative cocci (Image 2). The organism was identified by MALDI-TOF as Aggregatibacter aphrophilus. Antibiotic susceptibility testing showed susceptibility to augmentin, ampicillin, ceftriaxone, and levofloxacin.

Image 1. Growth on anaerobic chocolate plate.
Image 2. Gram stain from anaerobic culture showing gram negative cocci.

Discussion

Aggregatibacter aphrophilus is a gram negative coccobacillus that requires 5% CO2 and grows best on blood agar. It is oxidase negative and catalase negative. It is categorized as a HACEK organism, being a cause of culture-negative endocarditis. It is considered normal oral flora, and dental procedures can be a source of infection. Aggregatibacter endocarditis can cause a positive P-ANCA and be misdiagnosed as a vasculitis. It has also been reported as causes of sacroiliitis, bartholinitis, endophthalmitis, and brain abscesses. Treatment is generally ceftriaxone for 8 weeks. Identification is by biochemical methods or MALDI-TOF. Broad range PCR (br-PCR) has also been described, which targets a highly-conserved region of 16S rDNA, and then compares the sequences to database sequences.

The patient was given cefazolin, and his temperature downtrended. He was discharged prior to results but placed on oral augmentin. After susceptibility testing, infectious disease was consulted and he was placed on ceftriaxone for 8 weeks. He continued to improve and subsequent cultures were negative.

References

  1. Ratnayake L, Olver WJ, Fardon T. Aggregatibacter aphrophilus in a patient with recurrent empyema: a case report. J Med Case Rep. 2011;5:448. Published 2011 Sep 12. doi:10.1186/1752-1947-5-448
  2. Hirano K, Tokui T, Inagaki M, Fujii T, Maze Y, Toyoshima H. Aggregatibacter aphrophilus infective endocarditis confirmed by broad-range PCR diagnosis: A case report. Int J Surg Case Rep. 2017;31:150–153. doi:10.1016/j.ijscr.2017.01.041

-Jonathan Wilcock, MD is a 1st year anatomic and clinical pathology resident at the University of Vermont Medical Center.

-Christi Wojewoda, MD, is the Director of Clinical Microbiology at the University of Vermont Medical Center and an Associate Professor at the University of Vermont.

Hematopathology Case Study: A 36 Year Old Woman with an Incidental Neck Mass

Case History

A 36 year old female underwent thyroidectomy for multinodular goitre that led to the fortuitous discovery of a neck mass. The neck mass specimen submitted comprised two lymph nodes measuring 2.2 cm and 1.3 cm in the greatest dimensions, with a fleshy tan cut surface.

Biopsy Findings

H&E stained sections revealed numerous non-necrotizing granulomas effacing and replacing normal lymph node architecture. These consisted of pale epithelioid histiocytes and Langhans type of giant cells. The granulomas lacked a peripheral rim of lymphocytes. AFB and GMS stains were negative for microorganisms

Diagnosis

A diagnosis of non-necrotizing granulomatous lymphadenitis was rendered noting that in the correct clinical context the findings could represent sarcoidosis.

Discussion

Granulomatous inflammation is a special type of chronic inflammatory response characterised by the formation of discrete collections of histiocytes called granulomas. Activated histiocytes appear as epithelioid cells with round to oval nuclei, often with irregular contours and abundant granular eosinophilic cytoplasm with indistinct cell borders. They may coalesce to form multinucleated giant cells. When found in the lymph node, the reaction pattern is called granulomatous lymphadenitis. It can be caused by a variety of different conditions, and therefore, requires thorough workup to come to a conclusive diagnosis.

On the basis of presence or absence of necrosis, granulomatous lymphadenitis can be classified as necrotizing or non-necrotizing. Additionally, the presence of an abscess, usually central, indicates a suppurative lymphadenitis.

Non-necrotizing granulomatous lymphadenitis:

Sarcoidosis lymphadenitis is the prototype of non-necrotizing granulomatous lymphadenitis. It shows the presence of discrete granulomas without a peripheral rim of lymphocytes, called “naked granulomas”. The early phase shows follicular hyperplasia and sinus histiocytosis, followed by appearance of epithelioid cell nodules toward the end of this phase. The peak phase shows well-demarcated granulomas composed of epithelioid cells with scattered multinucleated giant cells observed throughout the lymph node. Granulomas may occasionally coalesce. In the late phase, increased collagen fibers result in fibrosis and hyalinization. There are no neutrophils and it is uncommon to find small foci of central necrosis. Numerous inclusions such as asteroid, Schaumann, or Hamazaki-Wesenberg bodies can be seen. In this case, we observed well-demarcated granulomas throughout the lymph node, typical of the peak phase without any caseous necrosis or suppuration.

Other causes of granulomatous lymphadenitis can be ruled out as follows.

Sarcoid-like lymphadenitis: It shows a similar pattern of non-necrotizing lymphadenitis like sarcoidosis. However, classically sarcoid like reaction shows scattered small epithelioid granulomas with sparsely arranged epithelioid cells. The border of the granulomas is usually obscure. The CD4:CD8 ratio ranges from 0.8 to 2.25 while in sarcoidosis, it is >3.5. These findings help distinguish sarcoid-like lymphadenitis from sarcoidosis.

Sarcoid-like adenitis may be seen in numerous conditions such as carcinoma, Toxoplasmosis, fungal infections, tuberculosis, immunocompromised states, pneumoconiosis etc. The fact that tuberculosis and fungal infections can present with a non-necrotizing granulomatous lymphadenitis highlights the importance of performing fungal (PAS & GMS) and AFB (Ziehl Neelson) stains in non-necrotizing lymphadenitis as well. In this case, the granulomas had distinct borders, numerous epithelioid cells, no organisms were identified on special stains, nor was there any history of immune compromise; ruling out a sarcoid-like reaction.

Berylliosis: The lymph node picture in Berylliosis is identical to that of sarcoidosis. We may even see asteroid bodies or Schaumann bodies. A diagnosis can be established by eliciting a history of chronic exposure to Beryllium. Beryllium lymphocyte proliferation test (BeLPT) is a test that measures Beryllium sensitization and is very specific for Beryllium exposure. There was no known history of exposure to Beryllium in this case.

Toxoplasmosis: A classic triad of follicular hyperplasia, small granulomas composed of epithelioid cells within and around hyperplastic follicles and, monocytoid B cell hyperplasia, is observed in toxoplasmosis lymphadenitis. This case did not show follicular hyperplasia, ruling out toxoplasmosis.

Necrotizing granulomatous lymphadenitis

Even though we did not find any necrosis in this case, yet, it is worthwhile to review briefly the various causes of necrotizing lymphadenitis.

  • Non-suppurative

Tuberculosis: Histology of a tuberculous lymph node is characterised by central caseous necrosis surrounded by an epithelioid cell layer. The outermost layer is comprised of lymphocytes and fibrosis. Plasma cells are not observed. Diagnosis can be established by performing an AFB stain that demonstrates acid fast rod shaped bacteria in the areas of necrosis. Organisms can also be detected by PCR.

BCG lymphadenitis: About 0.7 to 2.3% of BCG vaccinated children may develop BCG lymphadenitis that is smaller than tuberculous lymphadenitis. Early phase shows follicular hyperplasia and sinus histiocytosis. Later, there is development of micronodules of epithelioid granulomas without necrosis and epithelioid cell granulomas with central caseous necrosis. Langhans giant cells are rare.

Fungal infections: Fungal infections by Histoplasma, Cryptococcus, coccidiodomycosis, pneumocystis may also cause a necrotizing granulomatous inflammation. There are numerous neutrophils, and fungal structures can be seen. GMS and PAS can be used in cases where it is difficult to the find the fungal elements on H&E.

  • Suppurative

Tularemia: There are three forms of histological changes, Abscess form, showing abscess with central necrosis and mononuclear cells, Abscess-granulomatous form with granulomas with central necrosis, which form large lesions with central abscesses, and granulomatous form with caseating necrosis at the centre of the granulomas.

Cat Scratch disease: Similar to tularemia, there are three phases of histologic presentation, an early phase of follicular hyperplasia, intermediate phase of microabscess, and a late phase of granulomatous inflammation. Monocytoid B cell clusters are observed close to the abscess.

Conclusion

Sarcoidosis is usually diagnosed by excluding other causes of granulomatous inflammation, as we did in this case. Characteristic non-necrotizing, discrete granulomas were seen throughout the lymph node. The age of the patient and female gender epidemiologically support the diagnosis. This case reflects an example work up of a granulomatous lymphadenitis that is a morphologic presentation of myriad diseases.

-Swati Bhardwaj, MD has a special interest in surgical pathology and hematopathology. Follow her on Twitter at @Bhardwaj_swat.

Kamran M. Mirza, MD, PhD, MLS(ASCP)CM is an Assistant Professor of Pathology and Medical Education at Loyola University Health System. A past top 5 honoree in ASCP’s Forty Under 40, Dr. Mirza was named to The Pathologist’s Power List of 2018. Follow him on twitter @kmirza.

Surgical Pathology Case Study: A 43 Year Old Female with a Lung Nodule Noted on Imaging Following Chest Congestion

Case History

The patient is a 43 year old woman who experienced chest congestion and presented to her local physicians office. A chest X-ray was ordered and demonstrated a lung abnormality. A follow-up CT scan confirmed a 1.9 cm smoothly marginated nodule in the upper lobe with no adenopathy and a normal liver and adrenal glands. The nodule was mildly hypermetabolic on PET scan. A bronchoscopy was performed, which was non-diagnostic. Two subsequent CT scans demonstrated no change in the size of the nodule. Overall, the patient feels well and denies cough, hemoptysis, dyspnea on exertion, and weight loss. Due to the suspicion of cancer, the patient has decided to undergo a lung lobectomy.

Diagnosis

Received in the Surgical Pathology lab for intraoperative consultation is a 30.0 x 7.2 x 2.2 cm lung lobectomy specimen. There is an attached 6.2 cm staple line, which is removed and the subjacent resection margin is inked blue. The entire pleural surface is inked black. The specimen is sectioned revealing a 2.1 x 1.7 x 1.0 cm white-tan, firm, round nodule that is 0.5 cm from the blue inked resection margin and 0.2 cm from the black inked pleural surface. The remainder of the specimen is composed of red-tan, spongy, grossly unremarkable lung parenchyma without nodules or other lesions. Photographs of the specimen are taken (Figure 1). A representative section of the nodule is submitted for frozen section and read out as “diagnosis deferred”. Representative sections of the specimen are submitted as follows:

A1FS:   Frozen section remnant

A2-A7:   Nodule, entirely submitted

A8-A10:   Grossly unremarkable lung parenchyma

Immunohistochemical stains show the epithelial cells in the lesion to be positive for CK7, TTF-1, and surfactant proteins A and B which supports these cells to be type 2 pneumocytes (all controls are appropriate). Based on the immunohistochemical stains and routine H&E slides, the case was signed out as a sclerosing pneumocytoma

Image 1. Gross presentation of the well-defined, round sclerosing pneumocytoma.

Discussion

Sclerosing pneumocytoma (SP) is a rare, benign pulmonary tumor that was first described in 1956 as a vascular tumor, but has since been found to be of primitive respiratory epithelium origin. In the past, SP has also been referred to as sclerosing hemangioma, pneumocytoma, and papillary pneumocytoma, but the 2015 World Health Organization classification of lung tumors states that the agreed upon term for this tumor should be a sclerosing pneumocytoma. SP is commonly seen in middle aged adults, with a female to male ratio of 5:1. There is no racial bias. Patients are usually asymptomatic, with the tumor incidentally found on screening chest radiographs. If the patient was to present with any symptoms, they would usually include a cough, hemoptysis and chest pain. Radiographically, SP appears as a solitary, well-defined, homogenous nodule along the periphery of the lung.

Grossly, most SPs appear as a solitary, firm, well-circumscribed, yellow-tan mass generally arising along the periphery of the lung. The majority of these tumors appear within the lung parenchyma, but there have been cases reported of endobronchial and pleural based SP tumors. Multifocal unilateral tumors and bilateral tumors are uncommon.

Histologically, SP consists of two epithelial cell types: surface cells and round cells. Surface cells are cuboidal, resembling type II pneumocytes, with finely stippled nuclear chromatin, indistinct nuclei, occasional nuclear grooves, and inclusions. The stromal round cells will have bland oval nuclei with coarse chromatin and eosinophilic cytoplasm (Figure 2). Both the surface cells and round cells will have a low mitotic rate, but can have moderate to marked nuclear atypia. Ciliated bronchial epithelium is often identified in the tumor. There are four architectural patterns identified within SP: papillary, sclerotic, solid and hemorrhagic, with over 90% of SPs displaying three of the patterns, and all of the tumors containing at least two of the patterns.

  • Papillary pattern: Complex papillae composed of surface cells covering a stroma of round cells
  • Sclerotic pattern: Papillae containing hyalinized collagen, either in solid areas or along the periphery of hemorrhagic areas (Figure 3)
  • Solid pattern: Sheets of round cells bordered by surface cells
  • Hemorrhagic pattern: Large blood filled spaces
Image 2. Photomicrograph demonstrating the cuboidal surface cells and round stromal cells.
Image 3. Photomicrograph of the papillary and sclerotic architectural patterns.

Immunohistochemical stains can be helpful in the diagnosis of SP, with both the surface cells and round cells exhibiting expression of thyroid transcription factor 1 (TTF-1) and epithelial membrane antigen (EMA). It should be noted that TTF-1 is also used for the diagnosis of pulmonary adenocarcinoma, increasing the risk of misdiagnosing SP. The surface cells will also express both pancytokeratin (AE1/AE3) and Napsin A, with the round cells being negative for AE1/AE3, but having a variable expression of cytokeratin 7 and the low molecular weight cytokeratin (CAM 5.2). Molecular pathology has demonstrated a frequent loss of heterozygosity at 5q, 10q and 9p, and an allelic loses at p16 in the surface and rounds cells. Although the immunohistochemical stains and molecular pathology results can be very helpful, diagnosis of a SP is still largely based on routine H&E slides showing the two epithelial cell types and four architectural patterns.

Electron microscopy will show abundant lamellar bodies similar to those in type II pneumocytes in the surface cells. Round cells will lack the lamellar bodies and instead will contain variably-sized electron-dense bodies that have been thought to represent the different stages of lamellar body maturation.

The differential diagnosis for SP includes a variety of benign and malignant neoplasms, which can be difficult to distinguish on cytology, small biopsies and intraoperative consultations. The cytologic features include moderate to high cellularity with a bloody background and foamy macrophages, occasional nuclear pleomorphism in the round cells, absent mitotic figures, and occasional necrosis with cholesterol clefts and calcifications. In the case of small biopsies, making a diagnosis of SP can be difficult if the papillary pattern is highly prevalent without one of the other three patterns present. With intraoperative consultations, the frozen section artifact can make it difficult to appreciate the two epithelial cell types or the four architectural patterns. The gross examination, as well as the radiographic findings of a well-circumscribed tumor can help point the Pathologist to favoring a benign neoplasm over a malignant one. The benign neoplasms that should be considered in the differential diagnosis include:

  • Clear cell tumor, which will have clear cells with scant stroma, thin-walled vessels and a strong expression of HMB-45
  • Pulmonary hamartoma, which will have a combination of cartilage, myxoid stroma, adipose tissue and trapped respiratory epithelium
  • Hemangiomas, which are rare in the lung, and will lack epithelial cells and contain either a cavernous or capillary morphology

The malignant neoplasms that should be considered in the differential diagnosis include:

  • Bronchioalveolar carcinoma, which can have a papillary pattern, but will not contain the two epithelial cell types and combination of the four architectural patterns
  • Metastatic papillary thyroid carcinoma, which is distinguished from SP by the presence of the characteristic Orphan Annie nuclei
  • Metastatic renal cell carcinoma, which will contain nuclear atypia and striking vascularity
  • Carcinoid, which will contain organoid and ribbon-like growth patterns

Currently, with the benign nature of SP, surgical excision is the preferred treatment choice to cure the patient. There have been cases reported of lymph node metastasis and recurrence, but neither of these appear to effect the prognosis. This just helps to highlight the need for a multidisciplinary approach to this benign tumor.

References

  1. Hisson E, Rao R. Pneumocytoma (sclerosing hemangioma), a Potential Pitfall. Diagn Cytopathol. 2017;45(8):744-749
  2. Keylock JB, Galvin JR, Franks TJ. Sclerosing Hemangioma of the Lung. Arch Pathol Lab Med. 2009;133(5):820-825.
  3. Travis WD, Brambilla E, Nicholson AG, et al. The 2015 World Health Organization Classification of Lung Tumors: Impact of Genetic, Clinical and Radiologic Advances Since the 2004 Classification. J Thorac Oncol. 2015;10(9):1243-1260.
  4. Wu R. Sclerosing Pneumocytoma (Sclerosing Hemangioma). Pathology Outlines. http://www.pathologyoutlines.com/topic/lungtumorsclerosingheman.html. Revised February 19, 2019. Accessed June 6, 2019.

-Cory Nash is a board certified Pathologists’ Assistant, specializing in surgical and gross pathology. He currently works as a Pathologists’ Assistant at the University of Chicago Medical Center. His job involves the macroscopic examination, dissection and tissue submission of surgical specimens, ranging from biopsies to multi-organ resections. Cory has a special interest in head and neck pathology, as well as bone and soft tissue pathology. Cory can be followed on twitter at @iplaywithorgans.

Microbiology Case Study: A 58 Year Old Female with Abdominal Pain

Clinical History

A 58 year old female with no significant past medical history presented her primary care physician with chief complaint of abdominal pain. She reported continued vague abdominal symptoms for the past two months, with intermittent diarrhea and increased flatulence. No recent travel history or significant exposures were identified. An ultrasound of the right upper quadrant was unremarkable and no gallstones were present. The patient was scheduled for a screening colonoscopy. A stool specimen was submitted to the microbiology laboratory for stool culture and ova & parasite exam.  

Laboratory Identification

Image 1. Trichrome stained fecal smear illustrating a binucleated trophozoite with fragmented karyosomal material from a stool ova & parasite exam.
Image 2. Additional trichrome fecal smear image highlighting both uninucleate and binucleate trophozoites that range in size from 5 to 15 um.

The findings from the ova and parasite exam were consistent with Dientamoeba fragilis, an intestinal flagellate. The stool culture was negative for Salmonella, Shigella, and Escherichia coli 0157:H7. Stool enzyme immunoassays were negative for Campylobacter spp.and Shiga toxin 1 and 2.

Discussion

Dientamoeba fragilis is an intestinal flagellate with worldwide distribution and causes asymptomatic and symptomatic infections, predominantly in small children. Symptoms of infection may include intermittent diarrhea, abdominal pain, anorexia, weight loss, and flatulence.  While the pathogenesis is not completely understood, transmission is thought to occur via the fecal oral route and it is hypothesized that the trophozoites are transmitted via the eggs of nematodes, Enterobius vermicularis and Ascaris lumbricoides, due to a higher incidence of co-infections between these organisms than expected.

In the laboratory, the diagnosis of D. fragilis is made by ova and parasite exam. The trophozoite resembles amebae and is typically 9-12 µm. Most trophozoites are binucleate with finely granular cytoplasm and the within the nuclei there are 4-8 fragments of karyosomal granules (Figure 1). Due to the fact that 30-40% of D. fragilis trophozoites are uninucleate (Figure 2) and they lack external flagella, they must be differentiated from Endolimax nana and Entamoeba hartmanni, which are both non-pathogenic amebae. Historically, no cyst phase was known for D. fragilis; however, recent studies have identified precyst forms or putative cysts. Permanently trichrome stained slides are essential to diagnosing D. fragilis infection, as the organism is hard to detect in concentrated smears.

Since our patient was symptomatic, she was treated with iodoquinol, the drug of choice for D. fragilis infections. Her symptoms resolved and colonoscopy did not reveal additional pathology.  

-Debbie Walley, MD, is a 4th year Anatomic and Clinical Pathology chief resident at the University of Mississippi Medical Center. 

-Lisa Stempak, MD, is an Assistant Professor of Pathology at the University of Mississippi Medical Center in Jackson, MS. She is certified by the American Board of Pathology in Anatomic and Clinical Pathology as well as Medical Microbiology. She is the Director of Clinical Pathology as well as the Microbiology and Serology Laboratories. Her interests include infectious disease histology, process and quality improvement, and resident education.