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Samples of a 5-day-old Hispanic infant male (birthweight >3.00kg) were sent to the Biochemical Genetics Laboratory of Mayo Clinic to confirm an abnormal newborn screen result consisting of elevated short- to long-chain acylcarnitines (C4–C18). Plasma acylcarnitine analysis confirmed the abnormal elevations of C4-C14 acylcarnitines, and further revealed elevated glutaryl carnitine concentrations (Figure 1). In addition, urinary organic acid and acylglycine profiles indicated elevated excretions of various organic acids/glycine conjugates, of which those pertinent to this case are annotated in Figure 2. Whole exome sequencing (WES) performed in confirmation of the biochemical results revealed two compound heterozygous variants (classified as variants of unknown significance) in FLAD1. Although this patient is below the 7th percentile for growth/weight of infants his age and has visible atypical lipid accumulation in his arms, legs, torso, and face, he remains relatively asymptomatic at the current age of 12 months.
The correct answer is ...
Secondary glutaric acidemia type II
1. Ethylmalonic encephalopathy
Ethylmalonic encephalopathy is characterized as an autosomal recessive disorder caused by pathogenic variants in a mitochondrial persulfide dioxygenase encoding gene (ETHE1) required for H2S catabolism. Impaired regulation of this mechanism with the subsequent accumulation of H2S inhibits various biochemical enzymes such as cytochrome C oxidase (electron transport chain; mitochondrial dysfunction), short branched-chain acyl-CoA dehydrogenase (isoleucine metabolism; short branched-chain acyl-CoA dehydrogenase deficiency), and short-chain acyl-CoA dehydrogenase (β-oxidation; short-chain acyl-CoA dehydrogenase deficiency). Considering this, patients with ethylmalonic encephalopathy typically present with elevated concentrations of C4 and C5 acylcarnitines, ethylmalonic acid, methylsuccinic acid, isobutyrylglycine, 2-methylbutyrylglycine, and lactic acidemia.
Although strikingly similar to the presented case, ethylmalonic encephalopathy is unlikely to account for some of the other biochemical findings such as medium- and long-chain acylcarnitines, 3-hydroxyglutaric acid, and glutarylcarnitine associated with the present case. Clinically, patients with ethylmalonic encephalopathy characteristically present with developmental delay, seizures, petechiae, and orthostatic acrocyanosis early in infancy, which does not correlate with the current case.
2. Medium-chain acyl-CoA dehydrogenase deficiency (MCADD)
MCADD is a mitochondrial β-oxidation disorder that impairs the catabolism of medium-chain fatty acids (C6-C10) and the subsequent production of ketone bodies, due to biallelic pathogenic variants in the ACADM gene. These patients typically present with seizures, vomiting, lethargy, and hypoketotic hypoglycemia during fasting periods, which may lead to sudden/unexpected patient death. A potential biochemical diagnosis of MCADD should be considered in patients with elevated medium-chain plasma acylcarnitines, especially when arranged in a characteristic C6<C8>C10 pattern. Confirmatory biochemical observations include elevated hexanoylglycine, suberylglycine, and dicarboxylic aciduria.
A diagnosis of MCADD is excluded, as it doesn’t account for the elevated concentrations of short- and long-chain acylcarnitines, or the prominent excretions of ethylmalonic and methylsuccinic acids. Furthermore, no ACADM pathogenic variants were annotated during WES.
3. Glutaric acidemia type II (GA II)/multiple dehydrogenase deficiency
Patients with GA II were originally characterized as possessing biallelic pathogenic variants in flavoprotein related genes (ETF, ETFB, and/or ETFDH) responsible for transferring electrons, generated via numerous dehydrogenase reactions, to the electron transport chain. Impaired functionality of these flavoproteins thus concomitantly hamper dehydrogenase enzymes associated with fatty acid (β-oxidation), branched-chain amino acid, lysine, and tryptophan catabolism. As such, these patients present with elevated short- to long-chain acylcarnitines (C4-C18), hydroxyacids, dicarboxylic acids, and acylglycine conjugates (C5–C10 most prominent). The clinical phenotype of GA II patients is relatively heterogeneous and may manifest at various ages. Late-onset GA II may clinically present at any time after the neonatal period and exhibit signs of hypotonia, lethargy, rhabdomyolysis, lipid storage myopathy, and severe metabolic acidosis during periods of fasting.
Biochemically, GA II seems to be a likely diagnosis in the presented case. However, ethylmalonic acid, methylsuccinic acid, and 2-methylbutyrylglycine excretions in the presented case appear to be more prominently elevated than usually observed in these patients and may be more fitting with ethylmalonic encephalopathy/ethylmalonic aciduria. As further indicated, no pathogenic genetic variants were annotated in any of the aforementioned genes associated with GA II.
4. Secondary GA II
More recently, it has been reported that patients with pathogenic variants in genes (SLC52A1–3, SLC25A32, and FLAD1) associated with riboflavin transport and catabolism, exhibit biochemical abnormalities resembling GA II and ethylmalonic aciduria. In this case, two variants of unknown significance were found in one of these genes, FLAD1. Mutations in FLAD1 lead to impaired functionality of flavin adenine dinucleotide synthase (FADS), and subsequently reduce the production of FAD. The latter, in turn, could not only directly affect a variety of FAD-dependent flavoproteins (i.e., electron transferring proteins, etc.), but also cause secondary hampering of multiple acyl-CoA dehydrogenase enzymes (among others), hence the biochemical profile similar to GA II and ethylmalonic aciduria. Clinically, these patients share similarities with both the aforementioned diseases, and could reportedly present at various ages. However, very heterogeneous presentations have been described in the patients with this rare disease, the most prominent being lipid storage myopathy, reoccurring respiratory infections, weight loss, as well as speech and swallowing difficulties.
Collectively considering all the biochemical findings, molecular variants in FLAD1, and the abnormal lipid accumulation in this otherwise slender infant, the most likely diagnosis is secondary GA II, due to FADS deficiency. Since this patient remains relatively asymptomatic to date, it is anticipated that the genetic variants present in this individual may allow for some residual FADS activity that meets the current biochemical demand. Whether this may be sustainable throughout the patient’s lifespan remains unknown.
Zinandré Stander, Ph.D.
Fellow, Clinical Biochemical Genetics
Mayo Clinic
Silvia Tortorelli, M.D., Ph.D.
Consultant, Biochemical Genetics
Mayo Clinic
Associate Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science
A 37-year-old woman with a past medical history of ureteral obstruction requires placement of a nephrostomy tube when one of her bilateral ureteral stents is encrusted. She is afebrile and without urinary symptoms. Collected urine is sent to the laboratory in an anaerobic vial for culture and Gram stain. The Gram stain is read as negative for organisms and WBCs. Seven days later, a portion of the anaerobic CDC plate appears as below: no growth on aerobic plates was seen.
The correct answer is ...
Mycoplasma hominis
Mycoplasma hominis belongs to a phylogenetic class called 'mollicutes,' which have no cell wall and thus are not seen on a Gram stain. This case illustrates that these bacteria can occasionally grow on routine culture media incubated in anaerobic or 5% CO2 conditions, but they more commonly require specialized media. The pictured colonies are very small, not present on aerobic plates, and have only grown after many (seven) days of incubation.
M. hominis is part of the normal genitourinary microbiota in many adults and typically does not cause any symptoms. The role of M. hominis in many human genitourinary diseases is uncertain for three main reasons:
1. It is often a normal commensal.
2. It is difficult to culture, although molecular assays are available.
3. It is usually isolated with other bacteria.
Because M. hominis lacks a cell wall, it is intrinsically resistant to cell wall-active antimicrobials. Susceptibility testing is standardized by the Clinical & Laboratory Standards Institute and can be performed at specialized laboratories like the Diagnostic Mycoplasma Laboratory at the University of Alabama at Birmingham.
Proteus mirabilis: P. mirabilis is a urease-producing organism that can cause calculi primarily made of struvite crystals, which could account for this patient’s stent encrustation. However, P. mirabilis is a gram-negative rod (GNR) that should readily Gram stain. Furthermore, P. mirabilis should present as large GNR colonies, perhaps with swarming motility, on routine culture media (e.g., MacConkey or EMB) earlier than seven days of incubation.
Escherichia coli: E. coli is the most common cause of urinary tract infection, but should be readily identified as a gram-negative rod upon Gram staining. Furthermore, E. coli should present as large GNR colonies on routine culture media (e.g., MacConkey or EMB) earlier than seven days of incubation.
Ureaplasma urealyticum: Like M. hominis, U. urealyticum is a mollicute that commonly colonizes the urinary tract and does not Gram stain because it lacks a cell wall. Unlike M. hominis, U. urealyticum is unable to grow on routine culture media and instead requires specialized media (e.g., 10B broth and A8 agar) to grow.
Peter Kundert, M.D., Ph.D.
Resident, Anatomic and Clinical Pathology
Mayo Clinic
@KundertPeter
Audrey Schuetz, M.D.
Consultant, Clinical Microbiology
Mayo Clinic
Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science
A 38-year-old woman with a medical history of smoking and hyperlipidemia presents with a one-month history of back pain and nausea. For the past day, she has had shortness of breath, decreased activity tolerance, chills, and lightheadedness. Chest sounds were clear, but she required oxygen supplementation. Her chest CT showed ground glass opacities with septal and interlobular thickening (so-called “crazy paving”-pattern). A bronchoalveolar lavage (BAL) was ordered and images of the fluid, as well as the cytology, are shown.
The correct answer is ...
Pulmonary alveolar proteinosis (PAP)
PAP is caused by the accumulation of lipoproteinaceous substance in the alveoli due to abnormal surfactant production or lack of surfactant clearing. While the etiology of the disease is not perfectly elucidated, the majority of cases are believed to be caused by autoimmune impairment of granulocyte-macrophage colony stimulating factor (GM-CSF), which allows for normal function and maturation of macrophages that would normally clear surfactant. The presence of anti-GM-CSF antibodies in both the serum and the exudate can be indicative of this autoimmunogenic etiology.
Presentation of PAP typically occurs in the third to sixth decade of life and is predominately seen in men. Symptoms are largely nonspecific, with progressive dyspnea being the most common, followed by cough that may or may not be productive. Patients may also experience other symptoms that include, but are not limited to, fatigue, weight loss, and fever. While imaging can show the characteristic “crazy paving” pattern in up to 85% of cases, it is important to note that this is not a specific finding and can be seen in other disease states like pulmonary edema and PCP pneumonia.
The diagnostic workup for PAP is generally initiated by a BAL. The appearance of the milky-appearing exudate alone may be helpful in diagnosing the disease in the appropriate clinical context. The cytologic evaluation can also make the diagnosis, with the presence of amorphous, PAS(+) material that is diastase resistant in a paucicellular background, as represented in the above images. A biopsy can, of course, be diagnostic and reveals alveoli diffusely filled with eosinophilic material. Exclusion of co-infections (e.g., Nocardia) is imperative.
With regard to the differential diagnosis, in PCP pneumonia we would anticipate seeing foamy exudate with distinct “dot-like” structures, which would be PAS(-) and GMS(+). Amyloidosis would also be PAS(-) and would show its characteristic green birefringence under cross-polarized light with Congo red staining.
The classic treatment is a “therapeutic BAL” that can result in almost instantaneous resolution of symptoms. Looking forward, however, are studies in which the supplementation of GM-CSF may be an effective treatment option, as it can supplement the ineffective or deficient GM-CSF in the affected patients. And, of course, given the autoimmune correlation in the majority of cases, more common treatments such as monoclonal antibodies and immunosuppressants may also be considered.
Philip Hurst, M.D.
Resident, Anatomic and Clinical Pathology
Mayo Clinic
@pathophil
Melanie Bois, M.D.
Consultant, Anatomic Pathology
Mayo Clinic
Assistant Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science
@MelanieBoisMD
A 63-year-old woman presented with persistent postmenopausal bleeding. MRI of the pelvis showed a 6.3 cm uterine mass. The patient underwent total hysterectomy and bilateral salpingo-oophorectomy with bilateral pelvic sentinel biopsies.
The correct answer is ...
Mesonephric-like adenocarcinoma
Mesonephric-like adenocarcinoma (MLA) is a recently recognized aggressive adenocarcinoma variant of the uterine corpus and ovary. Patients often present at advanced stage. MLA has a propensity to metastasize to the lung.
Morphologically, MLA exhibits heterogeneous architectural growth, including a mixture of tubular, papillary and slit-like patterns. Tubules frequently contain intraluminal eosinophilic secretions. The nuclear features are low grade and can resemble papillary thyroid carcinoma by the presence of angulated nuclei and occasional nuclear grooves.
The typical MLA immunophenotype consists of expression of TTF1 and GATA3 with limited to no expression of ER and PR. This immunoprofile assists in their distinction from endometrioid carcinoma.
MLA of the uterine corpus has considerable morphologic and immunohistochemical overlap with mesonephric adenocarcinoma of the cervix, with a significant distinguishing feature being MLA lacks associated mesonephric remnants or hyperplasia (Wolffian structures located predominantly in the para-ovarian region and deep in the lateral aspect of the cervix).
MLA and mesonephric adenocarcinoma exhibit similar molecular aberrations consisting of KRAS mutations, gain of chromosome 1q, 10 and 12, and lack of PTEN alterations. PIK3CA mutations are also found in MLA, providing evidence of both mesonephric and Mullerian differentiation.
Farah Baban, M.B., Ch.B.
Fellow, Anatomic and Clinical Pathology
Mayo Clinic
@Baban.Farah
J. Kenneth Schoolmeester, M.D.
Consultant, Anatomic Pathology
Mayo Clinic
Assistant Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science
A 27-year-old women presented with abdominal pain and early satiety. Subsequent imaging demonstrated a 12 cm right ovarian mass. Her admission labs were all within the normal range, except for her serum calcium, which was modestly elevated. The patient underwent a unilateral salpingo-oophorectomy with salpingectomy with the following findings:
The correct answer is ...
Small cell carcinoma of the ovary, hypercalcemic type
Small cell carcinoma of the ovary, hypercalcemic type (SSCOHT), is a rare but aggressive ovarian tumor. It affects younger women, with an average age of diagnosis of 24. While the majority of patients will have hypercalcemia (>60%), a minority of patients will not have significant hypercalcemia prior to diagnosis. Symptomatic hypercalcemia is relatively uncommon. The clinical outcomes for SSCOHT are poor, with survival of early (stage I) disease as low as 33% and overall survival estimated to be 10% to 20%.
Histologically, these tumors can have a range of morphologies, including diffuse, trabecular, and nested architectures. So called ‘follicle-like’ spaces are often present, although they will be absent in some cases, such was the case in the current patient. Cytologically, the constitutive cells are usually small with scant cytoplasm, vesicular chromatin, and small nucleoli. A population of larger cells is also often present, characterized by abundant cytoplasm and with an eccentrically placed nuclei and prominent nucleoli. Rarely, large cells will make up the majority of the cellularity. Cells with prominent rhabdoid character are often present, which may be a useful finding. Mitotic activity is brisk, and necrosis is often present. A minority of cases will demonstrate mucinous epithelium, typically in the form of cystic spaces.
SCCOHT typically are immunoreactive for WT1, CD10, calretinin and EMA, and are negative for inhibin and SF1. Despite its name, neuroendocrine markers are positive in only a small minority of cases. Most significantly, the vast majority of SSCOHT show loss of SMARCA4. SMARCA4, also known as BRG1, is a member of SWItch/Sucrose Non-Fermentable (SWI/SNF) complex of proteins and is lost in many malignant neoplasms, including SMARCA4 deficient undifferentiated carcinomas of endometrium, SMARCA4 deficient thoracic sarcomas, and a subset of epithelioid sarcomas with retained SMARCB1/INI1. Loss of expression of this marker in combination with the above immunostains is useful in confirming the diagnosis.
The differential diagnosis for SSCOHT is broad, and immunohistochemical techniques greatly assist in narrowing the differential diagnosis. Granulosa cell tumors, both adult-type and juvenile-type, may show morphologic overlap and may present in a similar patient demographic. Markers of sex cord stromal differentiation, such as inhibin, SF1, and FOXL2 will be positive in granulosa cell tumors but would be negative in SSCOHT. A primary pulmonary-type small cell carcinoma would also be a consideration.
Morphologically, pulmonary-type small cell carcinoma will lack the characteristic follicle-like spaces, lack a large cell component, are often positive for TTF1, and more likely to be positive for neuroendocrine markers. They are usually negative for WT1 and demonstrate retained SMARCA4. Ovarian involvement by a hematopoietic neoplasm is sometimes a consideration due to morphologic similarities. Careful and thorough sampling should reveal a more conventionally epithelial component, and immunohistochemistry will readily differentiate these lesions. Metastatic tumors from extra-ovarian primaries should also be considered and excluded, including small cell carcinomas of pulmonary origin, endometrial carcinomas (including SMARCA4 deficient undifferentiated/dedifferentiated carcinomas), and melanoma, among others.
Mark Hopkins, M.D.
Fellow, Surgical Pathology
Mayo Clinic
Maryam Shahi, M.D.
Senior Associate Consultant, Anatomic Pathology
Mayo Clinic
Assistant Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science
A 22-year-old woman with no significant past medical history presented with a rapidly progressing right axillary violaceous plaque (measuring 2.0 X 1.4 cm) over a period of three weeks. Skin punch biopsy (Figure A) was performed, which demonstrated atypical CD3-positive T-cells as pictured in the figures. These cells co-expressed CD2, CD5 (partial), CD7 (very dim), CD56 (partial), TCR delta, TIA1 and granzyme B and lacked staining for CD4, CD8, CD20, CD30, CD138, TCR BF1, Cyclin D1, and EBER.
The correct answer is ...
Primary cutaneous gamma delta T-cell lymphoma
The clinical picture with the provided microscopic findings are highly suggestive of cutaneous lymphoproliferative disease. Diagnosis of a cutaneous T-cell lymphoma requires the presence of cytologically atypical T-cells, which often show an aberrant immunophenotype, sometimes requiring genetic corroboration to demonstrate clonality.
Morphologically, this case shows significant atypia. The neoplastic lymphoid cells are monomorphic, intermediate-sized, and have irregular nuclear contours. These cells infiltrate subcutaneous tissue and deep dermis and show adipocyte rimming (Figure B) and angiodestruction (Figure C). This pattern of lobular, panniculitis-like growth is typical for this lymphoma.
Immunophenotypically, our case shows partial expression of CD5, very dim CD7, partial CD56, dual CD4/CD8 negativity, and strong uniform expression of TCR delta with co-expression of cytotoxic markers TIA-1 and granzyme B. In combination, the above findings best support a diagnosis of primary cutaneous gamma delta T cell lymphoma.
In most cases, this rare T-cell cutaneous lymphoma has an aggressive clinical course with frequent mucosal and extracutaneous dissemination and the risk of hemophagocytic lymphohistiocytosis. Cases are often CD4-/CD8- but can occasionally express CD8. The 5-year overall survival is poor at 11% to 33%.
Mohammad Barouqa, M.D.
Fellow, Hematopathology
Mayo Clinic
@mohdbarouqa
Daniel Larson, M.D.
Senior Associate Consultant, Hematopathology
Mayo Clinic
