Lablogatory

Microbiology Case Study: Disseminated Disease Confirmed by Bone Marrow Biopsy in a Patient with HIV

Case History

A 35 year old female patient with a past medical history of uncontrolled HIV, retinitis caused by cytomegalovirus and recurrent colitis presented to the Emergency Department with body pain, fever, severe neutropenia, and diarrhea. CT scan revealed worsening sigmoid/rectal wall thickening. Patient also presented with esophageal candidiasis. Blood workup revealed that the patient had sickle cell disease (HBSC), anemia (Hgb 5.6 gm/dl) that required multiple transfusions, and elevated white blood cell count (up to 17,000). The patient also had leukopenia (neutropenia and lymphopenia), which, in addition to the anemia without hemolysis or bone marrow compensation and CD4 count <50, led to strong suspicious of disseminated mycobacteria infection. A bone marrow biopsy was performed and AFB staining revealed loose granulomas and numerous acid-fast bacilli seen. Culture of the bone marrow grew out acid-fast bacilli further identified as Mycobacterium avium complex (MAC).

Discussion

Mycobacterium avium complex (MAC) is made up of several nontuberculosis mycobacterial (NTM) species that require genetic testing to be speciated.¹ MAC is predominantly made of the slow-growers mycobacteria (SGM) such as M. avium, M. intracellulare, M. chimaera, and M. colombiense.^2,3 Most species of nontuberculosis mycobacteria are found in environmental sources. The MAC organisms are found throughout the environment, particularly in the soil and water, mainly in the Southeast of the United States.¹Human diseases are most likely from exposure to environmental sources either through direct inhalation, implantation or indirect consumption or contamination food or water. MAC is considered the most commonly encountered group of slow growers.

The MAC cause pulmonary disease that is clinically similar to tuberculosis, mostly in immunocompromised patients with CD4 cell counts less than 200/μL, such as those with HIV/AIDS. They are the most frequent bacterial cause of illness in patients with HIV/AIDS and immunosuppression.^1,4 MAC is also the most common nontuberculosis mycobacterial species responsible for cervical lymphadenitis in children. Additionally, hypersensitivity pneumonitis-like symptoms can occur which were initially thought to be an allergic reaction only, but current studies suggests infection and inflammation. Traditionally, MAC cause chronic respiratory disease, populations such as middle-aged male smokers and postmenopausal females with bronchiectasis (also known as Lady Windermere syndrome).

Diagnostic testing for pulmonary infection caused by MAC includes acid-fast bacillus (AFB) staining and culturing of the appropriate specimens. Respiratory specimens are the most commonly tested specimen type. If disseminated MAC (DMAC) infection is suspected, culture specimens should include blood and urine. Blood cultures are typically used to confirm the diagnosis of DMAC in an immunocompromised patient with clinical signs and symptoms ⁵. MAC can also be isolated from bodily fluids and other tissues, such as lymph nodes and bone marrow. If diarrhea is present, stool cultures can be collected. Skin lesions should be cultured if clinically warranted. To determine pulmonary involvement, imaging studies of the chest should be performed. Lymph node biopsy or complete lymph node excision is usually used to diagnose MAC lymphadenitis in children. Skin testing (MAC tuberculin test) has little value in establishing a diagnosis.⁶ Routine screening for MAC in respiratory or GI specimens is not recommended.

Organisms part of the MAC are not stained well by the dyes used in Gram stain, but instead are acid-fast positive. The ability of an organism to hold onto the carbol-fuchsin stain after being treated with a mixture of ethanol and hydrochloric acid is referred to as “acid-fast.” The high lipid content (around 60%) in mycobacteria’s cell wall makes them acid-fast. SGM require more than 7 days of incubation. Growth of M. avium species can be visualized in both LJ and 7H11 media ⁵. Colony morphology can be smooth or rough. Biochemical reactions to both niacin and nitrate reduction are negative. Upon growth, colonies can be identified using the MALDI-TOF mass spectrometry ⁶. However, depending on the database and technology used, reports from the MALDI-TOF may report MAC as M. avium complex or into the individual subspecies. Molecular techniques such as polymerase chain reaction or whole genome sequencing, as well as high-performance liquid chromatography, are required for species identification. Direct detection of nucleic acid in clinical specimens by PCR methods have been reported, although most tests are laboratory-developed and FDA-approved. Molecular technologies typically target the 16S rRNA gene, the 16S-23S internal transcribed spacer (ITS) region or the heat shock protein 65 (hsp65) gene. Prior to PCR, the AccuProbe test was the first commercial molecular assay for identification of mycobacteria by targeting 16S RNA ⁷. In Japan, an enzyme immunoassay (EIA) kit was used to detect serum IgA antibodies to MAC-specific glycopeptidolipid core antigen. This could be useful for serodiagnosis of pulmonary infections caused by the MAC. This EIA kit’s sensitivity and specificity have been reported to be 54-92% and 72-99%, respectively ⁸. Other serologic tests are also being investigated.

While this may not aid in the direct detection of MAC infection, a complete blood count (CBC) in DMAC patients frequently shows anemia and, on rare occasions, pancytopenia due to bone marrow suppression caused by the infection, though either leukocytosis or leukopenia may be present. Hypogammaglobulinemia may be another possibility ⁹. Patients with DMAC typically have elevated transaminase and alkaline phosphatase levels on liver function tests. An HIV test should be performed if pulmonary or disseminated MAC infection is suspected.

MAC is extremely resistant to antituberculosis medications, and a combination of up to six medications is often needed for effective treatment. The preferred medications at the moment are ciprofloxacin, rifabutin, ethambutol, or azithromycin combined with one or more of these other medications ⁴. For patients with HIV, azithromycin is currently advised as a preventative measure. Of note, preventive treatment of MAC colonization in asymptomatic patients is also not advised. The Clinical and Laboratory Standards Institute (CLSI) recommends performing antimicrobial susceptibility testing using broth microbroth dilution technique. Breakpoints for clarithromycin, amikacin, moxifloxacin, and linezolid are reported ¹⁰. Although ethambutol, rifampin, and rifbutin are useful, no official breakpoints are available as there are no strong correlation studies showing the relationship between minimal inhibitory concentrations (MIC) and clinical outcomes.

Figure 1. Acid-fast staining of the bone marrow aspirate revealed many acid-fast bacilli (left, 100X; right, 50X).

References

1. Akram SM, Attia FN. Mycobacterium avium Complex. StatPearls. Treasure Island (FL) ineligible companies. Disclosure: Fibi Attia declares no relevant financial relationships with ineligible companies.: StatPearls Publishing Copyright © 2023, StatPearls Publishing LLC.; 2023.

2. Miskoff JA, Chaudhri M. Mycobacterium Chimaera: A Rare Presentation. Cureus. 2018;10(6):e2750.

3. Murcia MI, Tortoli E, Menendez MC, Palenque E, Garcia MJ. Mycobacterium colombiense sp. nov., a novel member of the Mycobacterium avium complex and description of MAC-X as a new ITS genetic variant. International journal of systematic and evolutionary microbiology. 2006;56(Pt 9):2049-2054.

4. Kwon YS, Koh WJ, Daley CL. Treatment of Mycobacterium avium Complex Pulmonary Disease. Tuberculosis and respiratory diseases. 2019;82(1):15-26.

5. Hamed KA, Tillotson G. A narrative review of nontuberculous mycobacterial pulmonary disease: microbiology, epidemiology, diagnosis, and management challenges. Expert review of respiratory medicine. 2023:1-16.

6. Body BA, Beard MA, Slechta ES, et al. Evaluation of the Vitek MS v3.0 Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry System for Identification of Mycobacterium and Nocardia Species. Journal of clinical microbiology. 2018;56(6).

7. Ichiyama S, Iinuma Y, Yamori S, Hasegawa Y, Shimokata K, Nakashima N. Mycobacterium growth indicator tube testing in conjunction with the AccuProbe or the AMPLICOR-PCR assay for detecting and identifying mycobacteria from sputum samples. Journal of clinical microbiology. 1997;35(8):2022-2025.

8. Hernandez AG, Brunton AE, Ato M, et al. Use of Anti-Glycopeptidolipid-Core Antibodies Serology for Diagnosis and Monitoring of Mycobacterium avium Complex Pulmonary Disease in the United States. Open forum infectious diseases. 2022;9(11):ofac528.

9. Gordin FM, Cohn DL, Sullam PM, Schoenfelder JR, Wynne BA, Horsburgh CR, Jr. Early manifestations of disseminated Mycobacterium avium complex disease: a prospective evaluation. The Journal of infectious diseases. 1997;176(1):126-132.

10. CLSI. [Performance Standards for Susceptibility Testing of Mycobacteria, Nocardia spp., and Other Aerobic Actinomycetes, 1^st ed. CLSI M62. Clinical and Laboratory Standards Institute; 2018

-Dr. Abdelrahman Dabash is currently a PGY-2 pathology resident at George Washington University. He was born in Dakahlia, Egypt, and was raised in Al-Khobar, KSA. He attended the Faculty of Medicine at Cairo University, where he received his doctorate degree. He worked as an NGS analyst for 2 years prior to coming to GWU. His academic interests include Gastrointestinal pathology, hematopathology, and molecular pathology. In his spare time, he enjoys playing soccer, swimming, engaging in outdoor activities, and writing Arabic calligraphy. Dr. Dabash is pursuing AP/CP training.

-Rebecca Yee, PhD, D(ABMM), M(ASCP)^CM is the Chief of Microbiology, Director of Clinical Microbiology and Molecular Microbiology Laboratory at the George Washington University Hospital. Her interests include bacteriology, antimicrobial resistance, and development of infectious disease diagnostics.

Undetermined, Undetermined

I like to think most people who go into healthcare professions do so with the hope of helping others. For those of us who do autopsies, the greatest sense of reward comes when we can explain to someone how and why their loved one died. Inevitably, though, there will be situations where we need to accept what we don’t know – despite how disappointing it may be. In these situations, the most intellectually honest course of action is to issue a manner of “undetermined.”

Let’s recap briefly what we know about manners of death (see https://labmedicineblog.com/2022/11/25/please-dont-tell-me-i-died-of-cardiac-arrest/ for a more in-depth discussion). Manner of death describes the circumstances in which the cause of death is sustained, and there are five choices in most jurisdictions – natural, accident, homicide, suicide, or undetermined. Natural deaths are those due entirely to non-traumatic diseases (like cancer or coronary artery disease). Accidental deaths involve trauma or toxicity without an intention to harm. Homicide is death at the hands of another person, whereas suicide is death at one’s own hands. The final option is undetermined.

There are two main pathways by which we can arrive at an “undetermined” manner. There can either be 1) reasonable competing evidence between two manners of death, or 2) we may be unable to identify a cause of death due to loss or destruction of bodily tissue. Let’s look at some examples.

In the first pathway, consider an autopsy of a person with a single gunshot wound to the head. In a readily accessible region like the temple or beneath the chin, this wound could easily be self-inflicted. While this would be a “typical” location for a suicidal injury, such a wound could also be inflicted by another person. There are indicators we look for at autopsy which favor one scenario over the other. For example, most suicidal gunshot wounds (broadly speaking, of course) are contact wounds or intra-oral. A self-inflicted gunshot wound to the back of the head would be unusual, but (contrary to popular conception), not impossible depending on the firearm used. However, the same type of pattern could be elicited with another person holding a firearm to that individual’s head. We may examine the length of the firearm to determine if it’s possible for the decedent to have pulled the trigger themselves (keeping in mind that other items like a cane, coat hanger, or even the decedent’s toe, may have been used to depress the trigger). Similar questions can arise in autopsies of people who have fallen from height. There is no way an autopsy can tell with certainty whether an individual was pushed, fell accidentally, or left the edge of an elevated structure intentionally. The cause of death in both situations is undisputed – a gunshot wound in the first, and blunt force injuries in the second. This is why contextual information, like scene photographs and investigative records, is indispensable for forensic pathologists. Without context, we have no way to discern homicides, suicides, and accidents. Occasionally even with context, there can be competing narratives (one witness claims a gunshot wound was self-inflicted, while another claims it was inflicted by the first) or suspicious circumstances to cast doubt. Without clear cut evidence to support one story, the manner of undetermined is appropriate.

The second pathway by which we reach an undetermined manner is when extensive decomposition or other soft tissue loss (such as fire damage) interferes with our ability to determine a cause of death. Think of completely skeletal remains discovered in an abandoned building. Sometimes, indicators of potentially lethal injuries can still be identified – for example, a gunshot wound of the skull or knife marks on a rib. But, as the aphorism goes, “an absence of evidence isn’t necessarily evidence of absence” – a bullet or blade could be lethal while only striking soft tissue (especially in regions like the abdomen or neck). If we cannot rule out non-natural causes of death, the best choice for manner is “undetermined.”

An undetermined manner of death can understandably frustrate family members or law enforcement. I always try to explain that manner determinations are, as one of my mentors says, “written on paper and not in stone.” We reserve the right to change the ruling in the future if additional evidence comes to light. As forensics pathologists our primary responsibility is to speak honestly and truthfully, and sometimes that means admitting the limitations of our science.

-Alison Krywanczyk, MD, FASCP, is currently a Deputy Medical Examiner at the Cuyahoga County Medical Examiner’s Office.

Microbiology Case Study: A Case of Alpha-Gal Syndrome and Information on the Lone Star Tick

A 72 year old male presented to UVMMC in July, after being found unconscious and not breathing in his home. The patient presented with swelling of the throat and tongue, which had obstructed his airway. In addition to the swelling, the patient also presented with a hive-like rash along his upper torso and arms along with low blood pressure. The patient was successfully treated by an injection of epinephrine and asked about food allergies, as his clinical presentation was indicative of anaphylaxis. Having declared no food allergies, the patient was asked what he had eaten before the episode, noting that he had a beef burger for dinner hours earlier, which was not unusual for his diet. The attending physician noted the time between the man’s last meal and symptoms of anaphylaxis, which seemingly ruled out a food allergy. The patient was eventually discharged home, with recommendations to monitor his diet and return if symptoms resumed.

Two days later, the patient returned to UVMMC with coughing, shortness of breath, swelling of his tongue and throat, and heartburn. Once again, the patient was treated with injectable epinephrine, which alleviated his symptoms. When asked again about his diet, the man mentioned that hours earlier at dinner he had pork chops, which was also not unusual for his diet. Upon closer examination, a circular rash was observed on the patient’s right shoulder and the patient was tested for Lyme Disease. While awaiting the results of the test, the patient was asked about any exposures to ticks. Upon the mention of tick exposure, the man recalled seeing one a week prior crawling on his arm while he was watering his garden. Insisting that he did not feel a bite and quickly brushed the tick off of his arm, the man described the tick as being brown with a singular white dot on the center of its body. When the Lyme Disease test returned negative, the attending physician ordered a blood test, looking for specific antibodies to alpha-gal. The test returned positive, and the man was diagnosed with Alpha-Gal Syndrome (AGS) from exposure to a Lone Star Tick (Amblyomma americanum) bite. The patient was then referred to an allergist for symptom management.

*Figure 1.* Image of the rash discovered on the patient’s right shoulder

Lone star ticks (Amblyomma americanum) are aggressive human-biting ticks that actively seek out potential hosts through the use of CO₂ trails and vibrational movements.⁴ This strategy is a distinct behavior when compared to other tick species that commonly employ the ‘ambush strategy’ involving lying in wait for a potential host to pass by.^4,5 A complete life cycle for a lone star tick involves three distinct stages, including a larval, nymph and adult stage.³ While the bite of a larval tick is considered less dangerous due to it feeding for the first time and being less likely to have exposure to infected hosts, there is a risk that certain pathogens can be passed from the mother tick to the larvae.⁴ All three stages of the Lone Star tick’s life cycle require a blood meal from three different hosts, and all stages will feed on humans along with other vertebrate animals.³ These ticks live primarily in areas of woodlands where there is plenty of undergrowth and tall grasses.⁵

Due to changes in the climate, such as shorter, milder winters and an increased abundance of preferred hosts, the Lone Star tick has increased in both abundance and distribution over the last several decades.³ Despite these concerning trends, these ticks are commonly found throughout the eastern, southeastern, and south-central regions of the United States.³ Because Lyme Disease places such a huge burden on public health populations, the Lone Star tick is often overshadowed in public health messaging by black-legged ticks such as “deer” ticks (Ixodes scapularis) due to their Lyme-carrying abilities.⁴In contrast, the Lone Star tick is incapable of transmitting the spirochete that causes Lyme Disease (Borrelia burgdorferi)³, which is a reason why the patient’s blood test was negative for the pathogen in the current case.

Despite being incapable of carrying Lyme Disease, symptoms associated with a Lone Star tick bite may present similarly to that of Lyme Disease including the presence of a rash on the skin.^3,4 While similar, this rash is considered distinct from the rash observed in Lyme Disease and has been termed Southern Tick-Associated Rash Illness (STARI).³ While the specific etiologic agent has not yet been identified, the rash is often accompanied by fatigue, headache, fever, and muscle pains and will usually present within seven days of a tick bite.³ While no diagnostic test is available to distinguish STARI from Lyme disease, diagnosis is usually based on symptoms, geographic location, possibility of a tick bite, and the presentation of the rash which is typically a red circle expanding to around 8cm in diameter.³

Lone Star ticks can transmit a variety of bacterial and viral pathogens, but they are most commonly associated with Alpha-Gal Syndrome (AGS).^2,3,4,5 Alpha-Gal refers to the sugar molecule galactose-alpha 1,3-galactose, which is commonly found in most mammals except people, fish, reptiles, and birds.²The sugar molecule is found in meats (pork, beef, rabbit, lamb, venison, etc.), as well as in mammalian products such as gelatin, cow’s milk, or milk products.² Lone Star ticks transmit this sugar to humans by feeding on hosts and maintaining trace amounts of alpha-gal within their salivary glands, which is then injected into the next host.^2,4 In humans, the immune system reacts to alpha-gal in the bloodstream similarly to a foreign invader, initiating an IgE-mediated allergic response.⁴Symptoms will often vary between each individual but can include hives, nausea, vomiting, heartburn, dizziness or fainting, and anaphylaxis, among many other symptoms.^2,4

It is estimated that between 2010 and 2022, more than 110,000 people were suspected of having AGS, and diagnosis is usually confirmed by blood tests which look for specific antibodies to the sugar.² Interestingly, not every exposure to alpha-gal will result in an allergic reaction, and unlike food allergies where exposure can result in immediate reaction symptoms, it could take up to several hours after ingestion of an animal product containing alpha-gal for symptoms to appear in AGS patients.⁴Unfortunately, there is no treatment for AGS, but patients are typically managed by an allergist with recommendations of carrying an injectable epinephrine device, avoiding foods containing alpha-gal, taking antihistamines as needed, and monitoring or adjusting other medications which may be manufactured using gelatin capsules.^2,4

References

¹[Figure 1 Image]: ACP Internist. (n.d.). MKSAP Quiz: Evaluation for a Skin Eruption [website]. Accessed online on December 5^th 2023 from, https://acpinternist.org/archives/2016/10/mksap.htm

²CDC. (2023). Alpha-gal Syndrome [website]. Accessed online on November 17th 2023, from https://www.cdc.gov/ticks/alpha-gal/index.html

³CDC. (2018). Lone star tick a concern, but not for Lyme disease [website]. Accessed online on November 17th, 2023 from https://www.cdc.gov/stari/disease/index.html

⁴Kennedy, A. C., BCE1, & Marshall, E. (2021). Lone Star Ticks (Amblyomma americanum):: An Emerging Threat in Delaware. Delaware journal of public health, 7(1), 66–71. https://doi.org/10.32481/djph.2021.01.013

⁵Vermont Department of Health. (2023). Information on Ticks in Vermont [website]. Accessed online on November 17th, 2023 from https://www.healthvermont.gov/disease-control/tickborne-diseases/information-ticks-vermont

-Maggie King is a Masters student in the Department of Pathology and Laboratory Medicine at the University of Vermont.

-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.

Trust Your Gut

A 20 year old female patient referred herself to a surgical oncologist specializing in sarcomas after she presented to an outside hospital for a sudden onset of epigastric pain. The patient also reported a one-year history of decreased appetite without nausea, vomiting, or weight loss. The outside institution performed an abdominal ultrasound and identified a large nonvascular heterogenous masslike lesion in the left upper quadrant not definitively associated with the spleen or kidney. The mass measured 12.1 x 9.9 x 10.7 cm. The radiologist’s overall impression was a hematoma; however, a CT scan with contrast was recommended to further classify the lesion. Instead, an MRI was performed, and the same radiologist described the lesion as having a thick irregular enhancing rind with enhancing septations and central necrosis. With the lesion appearing distinct from adjacent organs, a retroperitoneal sarcoma was posited on imaging. Reviewing the outside imaging and clinical history, the surgical oncologist referred the patient to interventional radiology for an ultrasound-guided biopsy of the left-sided retroperitoneal mass.

When the cytologist arrived in the procedure room for the time-out, the radiologist informed her of the surgical oncologist’s and outside radiologist’s opinions of a retroperitoneal sarcoma. A 17-gauge coaxial needle was advanced into the peripheral and non-necrotic aspect of the retroperitoneal mass, and multiple 22-gauge fine needle aspirations were obtained and handed to the cytologist. She prepared two air-dried smears and two alcohol-fixed slides. The air-dried smears were stained in our Diff-Quik (DQ) set-up and deemed adequate. The pathologist’s immediate cytologic evaluation was “tumor cells present.”

Images 1-3: Retroperitoneum, Left-side, Ultrasound-guided FNA: DQ-stained smear.

The following morning, the cytologist primary screened the Papanicolaou-stained slides and H&E-stained cell block sections in addition to the DQ smears, with the former preparations presented below.

Images 4-7: Retroperitoneum, Left-side, Ultrasound-guided FNA. 4-5: Pap-stained smear; 6-7: H&E Cell Block section (400X).

The cytologist entered her results as positive for malignant cells with a note of “atypical cells in papillary fragments” and gave the case to the pathologist for the final interpretation. The pathologist reviewed the slides prior to ordering immunostains. He paused and thought, “there’s something about the morphology and her age… it just doesn’t make sense for this to be a retroperitoneal sarcoma. It doesn’t look like a sarcoma. The cells are just too round or ovoid, bland, and poorly cohesive, and the fibrovascular cores – I just don’t think this is a sarcoma. Maybe a melanoma? Or some type of renal tumor? The cytoplasmic vacuolization could suggest this, but the mass is distinct from the kidney, so it can’t be. The nuclear grooves are intriguing, almost like a papillary thyroid carcinoma. A neuroendocrine tumor is also possible, the delicate papillary fronds though… Hmm. But where would it be originating from? How could this be distinct from other organs in the abdominal cavity?” He hemmed and hawed, glancing over our list of in-house immunostains. With only nine pre-cut unstained sections associated with the three H&E cell block levels, the pathologist ordered additional unstained recuts. He knew this was going to be a challenge due to the discrepancy between the clinical history and the morphology.

With proper positive and negative controls, the tumor cells show positive staining for AE1/AE3, Cam 5.2, vimentin, CD99 (dot-like), CD56, beta catenin (nuclear), PR, AMACR, and SOX11, while negative staining for CK7, CK20, PAX-8, RCC, chromogranin, synaptophysin, GATA-3, EMA, GFAP, S100, calretinin, WT-1, E-cadherin, and p53 (wild type pattern). The proliferative index by Ki-67 is low at <1%.

Images 8-10: Retroperitoneum, Left-side, Ultrasound-guided FNA. 8: beta catenin (nuclear)-positive; 9: AMACR-positive; 10: SOX11-positive.

The combination of morphology with the extensive immunoprofile of the tumor is consistent with solid pseudopapillary neoplasm (SPN) of the pancreas.

Had there been any mention of the tumor involving or replacing the pancreas, this diagnosis and workup would have been much more straightforward. SPNs, albeit rare, account for 30% of tumors in women within their third or fourth decade of life.¹ This patient presented with the most common SPN symptoms of abdominal pain and early satiety, but the mass appearing extrapancreatic on imaging posed a diagnostic challenge, as extrapancreatic SPNs are rare.^2-3 Fortunately, SPNs are low-grade malignant neoplasms that respond well to surgical resection, and this patient is doing just fine after her distal pancreatectomy. In this case, both the patient and our pathologist listened to their guts with the patient pursuing advanced medical care for something much more complicated than a hematoma and the pathologist relying on his morphology expertise despite an odd clinical presentation.

References

La Rosa S, Bongiovanni M. Pancreatic solid pseudopapillary neoplasm: key pathologic and genetic features. Archives of Pathology & Laboratory Medicine. 2020;144(7):829-837. doi:10.5858/arpa.2019-0473-ra
Dinarvand P, Lai J. Solid pseudopapillary neoplasm of the pancreas: a rare entity with unique features. Archives of Pathology & Laboratory Medicine. 2017;141(7):990-995. doi:10.5858/arpa.2016-0322-rs
Cheuk W, Beavon I, Chui D, Chan JKC. Extrapancreatic solid pseudopapillary neoplasm. International Journal of Gynecological Pathology. 2011;30(6):539-543. doi:10.1097/pgp.0b013e31821724fb

-Taryn Waraksa-Deutsch, MS, SCT(ASCP)^CM, CT(IAC), has worked as a cytotechnologist at Fox Chase Cancer Center, in Philadelphia, Pennsylvania, since earning her master’s degree from Thomas Jefferson University in 2014. She is an ASCP board-certified Specialist in Cytotechnology with an additional certification by the International Academy of Cytology (IAC). She is also a 2020 ASCP 40 Under Forty Honoree.

Incidental Finding of Parasitic Infection in a 75 Year Old Male with Persistent Hiccups

Case History

A 75 year old man came to the Emergency Room because of intractable hiccups. He had a medical history of esophagitis, gastroesophageal reflux disease, gastric metaplasia diagnosed during a previous esophago-gastroduodenoscopy (EGD), and a significant episode of hiccups for several years. His esophagogastroduodenoscopy revealed diffuse edema and erythema on the duodenal mucosa. Histopathological examination of the duodenal biopsies (Figures 1-3) showed the presence of Strongyloides stercoralis within a few crypts of the duodenum, and adjacent eosinophil-rich inflammatory infiltrate within the mucosa. These findings provided an incidental finding of the parasite’s presence in the duodenal mucosa.

Figure 1. H&E stain of the biopsy at 10X

Figure 2. H&E stain of the biopsy at 400X

Figure 3. H&E stain of the biopsy at 400X

Discussion

Strongyloidiasis is a parasitic infection caused by the nematode Strongyloides – most commonly S. stercoralis. While it is commonly seen in tropical and subtropical regions, cases can also occur in temperate climates. Notably, our patient had a recent travel history to Jamaica, a known endemic region for Strongyloides infection.

The life cycle of Strongyloides stercoralis involves both free-living and parasitic stages. The infectious filariform larvae penetrate the human skin typically after contact with contaminated soil or exposure to infected fecal matter. Subsequently, they migrate to the lungs through the bloodstream, and eventually reach the small intestine, where they mature into adult worms. The adult worms reside in the duodenal and proximal jejunal mucosa, reproducing asexually by parthenogenesis. Some of the eggs hatch within the intestine, releasing rhabditiform larvae into the feces. It causes autoinfection by penetrating the intestinal wall or the perianal skin area.

The diagnosis of Strongyloides is typically accomplished by morphologic identification of larvae in the stool, duodenal aspirate, or sputum in disseminated cases. Strongyloides serologic testing is often performed in transplant patients who have a pertinent demographic and clinical history of potential exposure. The presence of eggs is rarely observed in the stool; therefore, microscopic examination of stool samples may have a lower sensitivity in uncomplicated infection with a low organism burden. In our case, stool samples were not collected for evaluation. Hyper-infection syndromes associated with disseminated Strongyloides could present as subclinical infection in patients under immunosuppression. As the larvae invade other organs, such as CNS, lungs, and blood stream, intestinal flora from the GI tract is carried along with the larvae, which causes super-infections, such as bacteremia and meningitis.

No FDA-cleared molecular testing is available for Strongyloides while some reference laboratories may offer laboratory-developed-tests. Therefore, the laboratory diagnosis frequently relies on the morphologic identification of the filariform larvae or eggs from clinical samples. In our case, the histopathological examination of the duodenal biopsies that were obtained to evaluate persistent hiccups revealed a significant eosinophil-rich inflammatory infiltrate within the mucosa, along with the presence of the larvae within the crypts. While hiccups can be due to various etiologies, including gastrointestinal disturbances and certain medications, and may not be directly related to parasitic infections, the diagnosis of Strongyloides in this case was purely incidental.

References

Gulwani, Hanni. “Strongyloides Stercoralis.” Pathology Outlines – Strongyloides Stercoralis, Aug. 2012, http://www.pathologyoutlines.com/topic/smallbowelstrongyloides.html.  
Carrada-Bravo, Teodoro. “Strongyloides Stercoralis: Vital Cycle, Clinical Manifestations, Epidemiology, Pathology and Treatment.” Revista Mexicana de Patolog, 1 Jan. 1970, http://www.medigraphic.com/cgi-bin/new/resumenI.cgi?IDARTICULO=16127.  
“Strongyloides Stercoralis.” RCPA, 2023, http://www.rcpa.edu.au/Manuals/RCPA-Manual/Clinical-Problems/S/Strongyloides-stercoralis.  
De la Cruz Mayhua, Juan Carlos, and Bisharah Rizvi. “Strongyloides Hyperinfection Causing Gastrointestinal Bleeding and Bacteremia in an Immunocompromised Patient.” Cureus, 24 June 2021, www.ncbi.nlm.nih.gov/pmc/articles/PMC8310433/.

-Inas Mukhtar, MD, is from Sudan and graduated medical school from University of Khartoum and started a pathology residency in Sudan before applying here to the US. She is currently PGY-2 at Montefiore Medical Center. Her hobbies include watching documentaries and spending time with friends and family.

-Phyu Thwe, Ph.D, D(ABMM), MLS(ASCP)^CMis Associate Director of Infectious Disease Testing Laboratory at Montefiore Medical Center, Bronx, NY. She completed her medical and public health microbiology fellowship in University of Texas Medical Branch (UTMB), Galveston, TX. Her interests includes appropriate test utilization, diagnostic stewardship, development of molecular infectious disease testing, and extrapulmonary tuberculosis.

Microbiology Case Study: Blastocystis in an Elderly Patient

A 68 year old female presented to our institution with complaints of lower right quadrant abdominal pain but an absence of nausea, vomiting, diarrhea and fever. The patient noted she thought she had a worm in her stool and submitted three stool samples for an ova and parasite exam. While the three stool samples were negative for worms and eggs, two of the three samples were positive for the organism seen below. The patient was treated with a regimen of metronidazole for ten days, and a repeat ova and parasite exam after treatment was negative for the parasite. The patient presented again three months later with similar complaints of abdominal pain, resulting in another ova and parasite exam which was again positive for the same organism.

Figure 1. *Blastocystis* spp. organism found in the most recent ova and parasite exam.

Figure 2. Another *Blastocystis* spp. organism found in the most recent ova and parasite exam (circled in red). The organism is in the vacuole form, with three nuclei visible on the periphery.

Originally discovered in 1911, Blastocystis species are anaerobic protozoan parasites commonly found in the human gastrointestinal tract.² In addition to being found in humans, Blastocystis spp. has also been found in a variety of animals, with at least nine different genotypes described in literature.⁴ While historically these organism have been referred to as Blastocystis hominis when found in humans, genotyping and molecular studies have revealed immense genetic diversity in organisms isolated from humans, resulting in a more general designation of Blastocystis spp.¹ These organisms have been observed worldwide, stemming from contaminated food, water and soil and transmitted in a fecal-oral route.² Recently, there has been much debate as to whether the parasite is truly pathogenic or commensal due to the presentation of both symptomatic and asymptomatic patients found carrying this parasite.⁴

Despite debate on the pathogenicity of Blastocystis spp., potential signs and symptoms from symptomatic patients have been documented. Such symptoms could include watery diarrhea, nausea, abdominal pain, bloating, excessive gas, loss of appetite, weight loss, and fatigue, among other symptoms.³ Diagnosis heavily relies on stool specimens using light microscopy to examine direct smears, with the vacuolar form of the parasite most commonly observed.¹ In addition to the vacuolar form, granular, amoeboid and cyst forms have also been described in literature.⁴ The parasite itself lacks a cell wall, but contains mitochondria, Golgi apparatuses, and both smooth and rough ER.² The vacuolar form, observed in Figure 1 and Figure 2, contains a large (5-40 mm) central vacuole-like body which compresses the nuclei to periphery of the cell.²

Treatment is typically not needed if the patient remains asymptomatic.³ However, for individuals who present with symptomatic infection, a regimen of metronidazole or tinidazole is recommended.³ It is important to note that due to the controversial nature of Blastocystis spp. as a pathogenic parasite, additional pathogens or causes of disease should first be ruled out prior to treatment.²

References:

¹CDC. (2019). Blastocystis sp. [online]. Accessed on October 24th, 2023 from https://www.cdc.gov/dpdx/blastocystis/index.html#:~:text=Blastocystis%20is%20a%20genetically%20diverse,been%20classified%20as%20a%20stramenopile.

²Leder, K. (2023). Blastocystis species [website]. Accessed online on October 26th, 2023 from https://www.uptodate.com/contents/blastocystis-species

³Mayo Clinic. (n.d.) Blastocystis Hominis [website]. Accessed online on October 24th 2023 from https://www.mayoclinic.org/diseases-conditions/blastocystis-hominis-infection/symptoms-causes/syc-20351205

⁴Wawrzyniak, I., Poirier, P., Viscogliosi, E., Dionigia, M., Texier, C., Delbac, F., & Alaoui, H. E. (2013). Blastocystis, an unrecognized parasite: an overview of pathogenesis and diagnosis. Therapeutic advances in infectious disease, 1(5), 167–178. https://doi.org/10.1177/2049936113504754

-Maggie King is a Masters Student in the Department of Pathology and Laboratory Medicine at The University of Vermont Larner College of Medicine.

-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.

Microbiology Case Study: Middle Aged Woman with Abdominal Pain

Case Description

A 59 year old female presented with a one-month history of generalized abdominal pain, difficulty breathing, and early satiety. She was previously seen at an external facility and found to have liver hypointensity and abdominal distension which were concerning for liver cancer. Social history included travel to Sacramento and California deserts for hiking in the last few months. CT of the abdomen and pelvis demonstrated extensive peritoneal carcinomatosis with no primary etiology and a right hepatic lobe focus. MRI revealed diffuse omental caking and peritoneal thickening consistent with peritoneal carcinomatosis. Hepatic lesions also seen on MRI were consistent with hemangiomas. Laboratory evaluation was positive for Hepatitis B virus infection along with elevated liver enzymes (AST and ALT), CA-125, and normal levels of CEA. Paracentesis was performed removing 3 liters of yellow, cloudy fluid. Cytology demonstrated benign mesothelial cells, macrophages, and mixed inflammatory cells with a lymphocytic predominance in the peritoneal fluid. No malignant cells were identified. Initial aerobic, anaerobic, mycobacterial, and fungal cultures were negative. Serum Cryptococcus neoformans antigen testing was also negative. Ultrasound-guided core biopsy of the omental mass was performed; however, tissue was not sent for culture. Histopathology revealed granulomatous inflammation with eosinophilic infiltrate (Figure 1). A singular spherule was highlighted by GMS stain (Figure 2). Fungal serology via complement fixation for Coccidioides was positive (titer: 1:8) while Blastomyces, Histoplasma, and Aspergillus serologies were negative. The patient was subsequently managed on fluconazole.

Figure 1. Core biopsy of omental tissue revealing granulomatous inflammation and diffuse eosinophilic infiltrate in a background of chronic inflammation, lymphocytes and fibrosis. Arrows indicate granuloma formation (H&E, 100x).

Figure 2. Core biopsy of omental tissue revealing a spherule with its thick wall containing endospores in a granuloma (GMS, 400x).

Discussion

Coccidioidomycosis is caused by the thermally dimorphic fungus Coccidioides that is endemic to the Southwestern United States, Mexico, and some regions in Central and South America. In endemic regions, farm workers, construction workers, military personnel, archaeologists, excavators, inmates, and correctional facility officers are most affected.¹ Individuals with significant exposure to the outdoors in endemic regions may be disproportionately affected. Multiple species of Coccidioides can cause disease and geographic distribution is changing²; however, the clinical presentation and approach to treatment of coccidioidomycosis are the same.

The pathophysiology of Coccidioides infection requires inhalation of arthroconidia which may result in pulmonary disease. Once inhaled, the organism forms an endospore-filled spherule in host tissue. Mature spherules rupture and disseminate endospores into surrounding tissue, which can then undergo additional replication cycles within the host. Symptomatic pulmonary disease appears to be related to fungal burden and usually presents with fever, fatigue, cough, pleurisy, and dyspnea, or less commonly peripheral eosinophilia and high erythrocyte sedimentation rate.³ Extrapulmonary disease, also known as disseminated coccidioidomycosis, is observed in <1% of cases. Extrapulmonary spread may develop in the meninges, skin, bone, or joints via hematogenous or lymphatic spread. Rarer sites of disease include the peritoneum, glandular tissue, visceral organs, bone marrow, reproductive organs, the pericardium, kidney, and the bladder.^3,4

The gold standard for diagnosis of coccidioidomycosis is culture; however, histopathology may yield definitive diagnosis, especially in cases where culture was not obtained. Following histopathological examination, polymerase chain reaction (PCR) from a variety of sources may also confirm the diagnosis, especially in cases without supporting clinical or laboratory data to suggest coccidioidomycosis.⁵ When culture is performed, Coccidioides is recovered in its environmental, or mold form. This phase is highly infectious and can pose a significant biosafety hazard; therefore, laboratory personnel should be appropriately notified when Coccidioides is suspected, especially since Coccidioides sp. can grow on solid media within the timeframe of conventional bacterial cultures.³ Serology can be used as an adjuvant test methodology in suspected cases. Serology may be performed on blood samples, CSF, joint, or pleural fluid and measured by immunodiffusion (ID), enzyme-linked immunoassay (EIA), or complement fixation (CF). ID uses gelatin as the medium in which antibodies are placed in one well and Coccidioides antigens are placed in another. Where the two meet through diffusion, a band will form from precipitated immune complexes. ID tests for both IgG and IgM antibodies.⁶ EIA, in contrast, uses enzyme-labeled antibodies to detect patientantibody complexed with bound antigen. CF incubates Coccidioides antigens with patient serum that has been heated (to inactivate endogenous complement). Following incubation, fresh, non-human serum containing complement is added. If immune complexes have formed between patient antibodies and the antigens, then the fresh serum complement will affix to those complexes and indicator red blood cells will remain intact when added. A positive result will have a suspension of intact red blood cells that can be titered. A negative test will result in lysed red blood cells. CF tests for IgG antibodies.⁶ Coccidioides antigen assays may be used to test the urine, blood, and CSF of patients with extrapulmonary disease. In immunocompromised hosts, serology may be negative, and coccidioidal antigen assay may be performed. Clinical management commonly involves long-term antifungal therapy with or without surgical intervention for high-risk individuals.⁴

References

“Epidemiology.” UC Davis Center for Valley Fever, https://health.ucdavis.edu/valley-fever/about-valley-fever/epidemiology/index.html. Accessed 6 July 2023.
Mazi PB, Sahrmann JM, Olsen MA, Coler-Reilly A, Rauseo AM, Pullen M, Zuniga-Moya JC, Powderly WG, Spec A. The Geographic Distribution of Dimorphic Mycoses in the United States for the Modern Era. Clin Infect Dis. 2023 Apr 3;76(7):1295-1301. doi: 10.1093/cid/ciac882. PMID: 36366776; PMCID: PMC10319749.
Proia L. Chapter 28: The Dimorphic Mycoses. In: Spec A, Escota G, Chrisler C, Davies B., eds. Comprehensive Review of Infectious Diseases. > ed. Elsevier; 2020: 411-423.
Johnson RH, Sharma R, Kuran R, Fong I, Heidari A. Coccidioidomycosis: a review. J Investig Med. 2021;69(2):316-323. doi:10.1136/jim-2020-001655.
Binnicker MJ, Buckwalter SP, Eisberner JJ, Stewart RA, McCullough AE, Wohlfiel SL, Wengenack NL. Detection of Coccidioides species in clinical specimens by real-time PCR. J Clin Microbiol. 2007 Jan;45(1):173-8. doi: 10.1128/JCM.01776-06.
“Explanation of Coccidioides Diagnostic Testing.” UC Davis Center for Valley Faver. https://health.ucdavis.edu/valley-fever/about-valley-fever/coccidioides-diagnostic-testing/index.html. Accessed October 26, 2023.

-Evelyn Ilori, MD, PhD is a third-year AP/CP resident at University of Texas Southwestern Medical Center in Dallas, Texas.

-Andrew Clark, PhD, D(ABMM) is an Assistant Professor at UT Southwestern Medical Center in the Department of Pathology, and Associate Director of the Clements University Hospital microbiology laboratory. He completed a CPEP-accredited postdoctoral fellowship in Medical and Public Health Microbiology at National Institutes of Health, and is interested in antimicrobial susceptibility and anaerobe pathophysiology.

-Clare McCormick-Baw, MD, PhD is an Assistant Professor of Clinical Microbiology at UT Southwestern in Dallas, Texas. She has a passion for teaching about laboratory medicine in general and the best uses of the microbiology lab in particular.

Three Safety Cultures Questions to ask Yourself, Your Staff and Your Leaders

Whether you are a newly graduated scientist or a seasoned individual starting at your fifth lab in your career, you might be surprised by the safety culture at the new facility. You could be so impressed by the safety culture at your new laboratory that you question how no one was seriously hurt at your former one. Or you could walk into the lab on your first day and immediately get a bad feeling in your gut. No matter how you feel on day one, two, or maybe day 32, just know that there are some things you can do to help understand your new perception of the culture. Any great piece of research starts with a question or two. Let’s examine some queries that can help you wrap your head around why some labs win, and others fall short when it comes to their safety culture.

First, let’s start with the why. When the safety culture does not look good, it is easy to assume that the deviant behaviors you witness are simply people taking advantage of the system. But not all bad behaviors are spawned from a desire to do harm. You need to find out what is influencing their unsafe behaviors. Most of the time, subpar safety behavior stems from a lack of understanding the consequences of unsafe actions. For example, some folks may not realize that handling their cell phone with gloved hands in the lab has the potential to transfer pathogens into the breakroom when they place that same phone on the table when they are eating their lunch. When the timing is right, you should have a conversation with the employee about what you saw and inquire if they are aware of the potential safety risks. You never know, you could discover that it was a topic skipped in safety training and you single-handedly just improved the quality of your safety training program!

The next question you should ask yourself is, are laboratory leadership aware of the safety issues present in their lab(s)? In most labs, the managers are often overburdened, spending most of their time chasing a schedule, trying to fill open positions, or putting out fires (figuratively we hope). Therefore, you should not assume that leadership is aware and allow unsafe practices to occur in the lab. Sometimes managers and supervisors are so hyper-focused on one thing, they might not be able to see a safety issue right in front of them. If you see unsafe habits, bring it up to lab leadership and share with them that your intentions are to avoid a potential harmful event from occurring. In some cases, managers are already aware of the situation and are trying to come up with solutions. Your conversation with them and perspectives about the safety concerns might be the missing piece that helps complete the puzzle they are trying to solve. So, you should feel comfortable bringing your concerns and be prepared to come with solutions to any problems you present to them.

Finally, ask yourself if the safety issues you see are isolated to a particular shift or certain individuals. Although it is the responsibility of laboratory leadership to champion the safety culture, it is up to the staff to feed and nurture its existence. When everyone works together, it is reflected positively in the safety culture, the audit results, and the injury and exposure reports. When gaps are present, there will be an increase in the negative indicators until the issues are identified and resolved. Instead of making assumptions about the safety culture of the entire lab, try to see where the gaps exist and then revisit the first question- why? It is a lot easier to coach a single individual that wears earbuds in the lab than the entire night shift crew that refuses to wear a lab coat until 5 minutes before the day shift supervisor appears. If that single person is the root of your safety concerns, don’t let their behavior go unchecked. As with negative attitudes in the department, poor safety habits can spread like wildfire. If a single individual’s behavior is not addressed, then others will soon follow suit. They will see that there are no repercussions to lax safety behaviors or worse, they will think nothing bad can come of cutting safety corners. Laboratory leaders and coworkers that normalize poor safety habits are only making the situation worse and damaging the safety culture of the lab while putting the entire staff at risk.

You should never assume a safety culture persists on its own. A good or bad safety culture is the sum of many different factors, and the reason behind the factors can be vast. So, before you are ready to write off a lab as unsafe, take the time to dig a bit deeper and find out what contributed to making the safety culture what it is today. If the lab has a great safety culture, find out why. The lab you are in today may not be the place at which you retire. Your path might lead to a different workplace that has an even worse safety culture than the one you left behind. By asking questions about what creates a great safety culture, you become equipped with the right tools and knowledge and will then be in a strong position to use what you know to improve the lives of others in your new lab.

-Jason P. Nagy, PhD, MLS(ASCP)^CMis a Lab Safety Coordinator for Sentara Healthcare, a hospital system with laboratories throughout Virginia and North Carolina. He is an experienced Technical Specialist with a background in biotechnology, molecular biology, clinical labs, and most recently, a focus in laboratory safety.

It’s What’s on the Inside That Counts: Uses of Radiography at Autopsy

In several previous blogs, I’ve mentioned the topic of post-mortem radiography (or “x-rays”). While postmortem CT scanning is a hot topic in the field, plain films are a tool which has been in widespread use for decades. Autopsy standards of the National Association of Medical Examiners require, at a minimum, radiographs be performed on all infants, gunshot wound victims, explosion victims, and charred or decomposed remains. Let’s examine the reasons for these requirements and look at a few specific examples.

All infants must get full body x-rays to check for acute or healing rib and long bone (extremity) fractures, which could be indicative of physical abuse. The extremities are not usually dissected in the course of a typical post-mortem examination, but fractures that can’t be attributed to birth trauma (especially in a pre-mobile child) are a concerning finding that needs further investigation and dissection.

Radiographs taken of gunshot wound victims document the presence and location of retained projectiles, all of which need to be recovered as evidence.

This x-ray of a gunshot wound victim shows two separate types of ammunition, seen as radio-opaque (white) in the image. The smaller, circular pieces are “birdshot” shotgun ammunition (blue arrows) while the larger pieces are traditional handgun ammunition (red arrow).

Radiographs are commonly performed in any type of penetrating trauma, including sharp force injury and explosive injuries, to identify retained foreign bodies (especially broken fragments of the blade, which may pose a risk to the pathologist). Similar to projectiles, these fragments need to be recovered as evidence. Radiographs can also document the presence of an air embolism, which can be missed at autopsy if special dissection techniques aren’t performed.

This individual had sharp force injuries to their neck, which injured large veins. The x-ray in this case was performed to see if any fragments of the weapon were still in the body, but also showed a large air embolism in the right atrium and ventricle (blue arrows), seen as radiolucency (gray/black). Open injuries to veins in the head and neck can cause air emboli as breathing creates negative intra-thoracic pressure, drawing air inward though any open channels.

In pedestrians who have been struck by motor vehicles, radiography is the first step in examining trauma. As mentioned earlier, the extremities aren’t typically dissected during a traditional autopsy – but in pedestrians, lower extremity fractures can document the site of initial impact, and the distance of the fracture from the foot may indicate the bumper height of the car.

*This x-ray of a pedestrian struck by a motor vehicle shows displaced fractures of the right femur, tibia, and fibula. Note the body bag zipper in the lower right corner.*

Fire victims may have extensive thermal injuries and charring which can hide evidence of other injuries, and radiographs can help identify them. Radiographs are also one step toward identifying the victim. Decomposed bodies must be x-rayed for similar reasons – external and internal soft tissue alterations make assessment for trauma more difficult, and radiographs may be needed to confidently identify the body.

In this x-ray of a decomposed person, dental fillings are easily visible (red arrow), which are typically unique for an individual and can be used for identification. Note the irregular shapes of the fillings, which are just as important as their location. Also note the air-fluid level in the cranial cavity (blue arrow), indicating complete liquefication of the brain.

On occasion, radiographs can be performed on individual organs to help define anatomy or previous surgical alterations. Especially in the heart, radiographs can demonstrate coronary artery or valve calcifications, surgical clips from bypass grafts, or other radiopaque prostheses. Knowing the location of devices before dissection gives the pathologist a better chance at preserving and evaluating important structures.

This heart was x-rayed after removal, as the decedent had a history of heart disease with prior interventions. Coronary stents (red arrows) are easily visible; in practice, they can be difficult to find on gross dissection in patients with heavily calcified arteries.

Not all offices can afford the installation (or maintenance) of CT scanners, but access to x-ray machines is more widespread. As we’ve seen here, x-rays are a versatile tool which can document injuries, help identify decedents, and direct the pathologist to perform special autopsy procedures which aren’t part of the daily routine.

-Alison Krywanczyk, MD, FASCP, is currently a Deputy Medical Examiner at the Cuyahoga County Medical Examiner’s Office.

As Laboratory Professionals, Are We Immune from Microaggressions?

Microaggression is a term first coined by Dr. Chester Pierce, a Harvard psychiatrist, in 1970, where he described the lived experiences of an African American man navigating in white predominant spaces. It took 37 years for the term microaggression to come into mainstream acceptance when it was re-introduced by Dr. Derald Sue in 2007, where he expanded the definition to include general disrespect, devaluation, and the exclusion of minorities.

I still remember my first frozen on-call as a PGY-1 AP/CP resident; I came to work at 6am to prep the gross lab. I was told to fill the big formalin container for the day ahead, log all the refrigerated patient specimens from overnight surgeries, ink to orient and then cut the specimens so they can be fixed. Then finally when I moved to change the water in the stain line, suddenly the tech walked in and asked, “Are you Muslim?” I paused and froze for a moment, didn’t have any words to answer his question. Then I said, “No!”, and he walked away. This incident set me back the whole day, because I was replaying it in my mind again and again, to see why he had asked this specific question to me. Maybe I should have answered differently. I didn’t know of the term, “microaggression” back in 2009.

Now I am equipped with this term and training my colleagues to respond in real-time to these types of instances, I wonder how empowered I would have been if I knew about microaggressions from med school/residency training. I would have been more confident in my interactions, rather than walking on eggshells.

When teaching bystander intervention, there are multiple mnemonics, so that you can quickly react to the situation rather than lack of response in the moment. The 5D’s from American Psychology Association that calls to action are: Distract, Delegate, Document, Delay and Direct. In JAMA Surgery, Dr. Nafisseh Warner introduced GRIT (Gather, Restate, Inquire, Talk It Out). In a learning environment to shift the power dynamics to the clinical learner, the visual below from Dr. Justin Bullock is helpful in breaking down each segment of bystander response: pre-brief, during and after the microaggression.

Reference: Bullock JL, O’Brien MT, Minhas PK, Fernandez A, Lupton KL, Hauer KE. No One Size Fits All: A Qualitative Study of Clerkship Medical Students’ Perceptions of Ideal Supervisor Responses to Microaggressions. *Acad Med*. 2021;96(11S):S71-S80.

When trainees come into laboratory spaces, it is essential to conduct a pre-briefsession to create a safe-learning environment. During orientation, talk to learners about the possibility of racism and bias. In medicine, the hierarchy is engrained and overemphasized within the system, so learners are hesitant to respond to microaggressions, report bias and racism. Therefore, taking the time to pre-brief will shift the power to learners to say something when being the target of racism and bias.

During microaggression, recognize and analyze it. Is it objectifying? Is it prejudice or negative attitude/stereotype? The usual response to microaggression is no response because you freeze, like I did in my scenario. The effective response is short and direct, most importantly respond in real-time. The degree of response will vary according to the situation and learner’s preference, sometimes the supervisor may bear witness or stop the interaction to create an opportunity for the trainee to exit.

After the microaggression, do not ignore what happened and move on. Supervisors should take the time to privately check-in with the learner and offer support, which is the most preferred approach. Team debriefs might invite an exhausting dialogue and may cause the target to relive the traumatic experience. Also, if there are instances of repeat microaggressions, propose the option for re-assignment.

To create a safe learning environment for our learners it is essential to prioritize psychological safety in our busy clinical services. Taking the time to pre-brief with learners, check-in and offer support to the target will promote a better work culture. Together we can support and empower each other to mitigate microaggressions at our workplaces.

-Deepti Reddi, MD, Assistant Professor of Pathology, Department of Laboratory Medicine and Pathology, UW School of Medicine; UW Medicine’s equity, diversity, and inclusion (EDI) peer trainer at the Office of Healthcare Equity (OHCE) and teaches Racism & Bias for Center for Learning & Innovation in Medical Education (CLIME) in Advanced Clinical Teaching Certificate program. She is also a 2021 ASCP 40 Under Forty Honoree.