Microbiology Case Study: A 59 Year Old Male with Acute Prostatitis

Clinical history

A 59 year old male with a past medical history of benign prostatic hypertrophy who presented to his primary care physician with complaints of dysuria, urgency, nocturia, and a weak stream. He was referred to a urologist who diagnosed him with acute prostatitis. The patient was given Flomax, a medication that relaxes the muscles in the prostate and bladder, and sulfamethoxazole-trimethoprim, an antibiotic for his possible bladder infection. The patient continued to have urinary symptoms despite antibiotics and multiple urinary cultures that were negative for growth of bacteria. Eventually the patient’s symptoms worsened and he presented to the emergency department with symptoms of fever (102.8ºF), bladder spasms, drenching night sweats, and painful urinary retention. His lab results showed a mild leukocytosis (15,500 cells /mm3) and increased leukocyte esterase, leukocytes, and erythrocytes in his urine. He was admitted and a pelvic computed tomography scan showed multiple prostatic abscesses. The infectious disease team ordered a urine fungal culture. The fungal stain showed broad-budding yeast forms (Image 1).

Image 1. Calcofluor white fungal stain highlighting brad-based budding yeast (100x oil immersion).

Laboratory identification

A cytospin Gram stain was made of the urine specimen which showed rare large yeast forms, numerous red blood cells and neutrophils (Image 2). These yeast forms are consistent with Blastomyces dermatitidis which are 10-15 µm in diameter and have thick contoured cell walls. The fungal culture grew a dirty white leathery mold on inhibitory mold agar after ten days. The lactophenol cotton blue adhesive tape preparation highlighted short to long conidiophores with large pear-shaped conidia at the tips of delicate conidiophores and septated hyphae. A positive urine antigen test supports the identification of the organism as Blastomyces dermatitidis.

Image 2. Large yeast with thick refractile walls seen on Gram stain. Flattened areas suggest recent separation.

Discussion

Blastomyces dermatitidis is a dimorphic fungus commonly found in the eastern half of the United States and Canada, specifically the Mississippi and Ohio River valleys, and Great Lakes Region. It typically grows as a mold in damp soil and decomposing vegetation; outdoor activities that disrupt the soil can increase the risk of infection. Infection occurs when the reproductive spores known as conidia are inhaled into the alveoli of the lungs. There the conidia transform into yeast and multiply causing the disease blastomycosis. Approximately 50% of those infected with Blastomyces are asymptomatic and clear the infection without trouble. For the other 50%, symptoms depend on the course of infection with a range of flu-like illness over a few days to chronic or severe illness that can last for months. Symptoms include fever, chills, headache, joint and muscle pains, shortness of breath, cough, night sweats, weight loss, and pleuritic chest pain. Twenty to forty percent of symptomatic patients will have disseminated disease. The common extra-pulmonary sites include skin, bone, urogenital, and the central nervous system, however, it has been reported in all organ systems.

In our case, the patient presented with prostatic blastomycosis which was quickly identified by calcofluor white fungal stain and urine antigen test. The patient was placed on itraconazole and a transurethral prostatic resection was performed. Histopathologic analysis showed necrotizing granulomas with fungal elements that are consistent with Blastomyces dermatitidis (Image 3). After a few days of treatment, the patient was discharged and is being followed on an outpatient basis.

Image 3. PAS stained tissue showing three budding yeast forms in a granuloma with extensive neutrophilic reaction.

– Joshua Wodskow, DO is a 1st year clinical and anatomic pathology resident at University of Chicago (NorthShore). Academically, Joshua has a particular interest in hematopathology and informatics. In his spare time, Joshua enjoys board games with his family and listening to podcasts.

-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: A Fishy Situation

Case History

The microbiology laboratory received a tissue specimen from a male patient in his 60s. The tissue was biopsied from the patient’s hand during surgery at an outside hospital, but no further clinical information was available. No organisms were observed on the Gram stain, fungal smear, or fluorochrome stain and no white blood cells were observed on the Gram stain. No organisms were recovered from the aerobic or anaerobic bacterial culture or fungal culture.

After 20 days of acid-fast bacilli (AFB) culture, yellow colonies grew on the Lowenstein Jensen (LJ) media and the 7H10 media (Image 1) and acid-fast bacilli were noted on the Kinyoun stain prepared from the colonies (Image 2).

Image 1. Yellow colonies on the 7H10 media.
Image 2. Kinyoun stain of the colonies demonstrating acid-fast bacilli. This is characteristic of Mycobacteria spp. Species-level identification is not possible based on morphology alone.

The ordering physician was contacted regarding the positive Kinyoun stain. Upon hearing that acid-fast bacilli grew in culture, the physician shared that he has a high suspicion for Mycobacterium marinum as this patient developed a wound infection after he cut his hand with a fish bone while cooking. MALDI-TOF identified this organism as Mycobacterium marinum, confirming the astute physician’s hypothesis. 

Mycobacterium marinum

Mycobacterium marinum is a slow growing non-tuberculous mycobacterium (NTM) that causes skin and soft tissue infections. M. marinum can be found in both fresh and saltwater environments and is commonly associated with contact with fish, fish tanks, marine shells, and non-chlorinated swimming pools, earning the names “swimming pool granuloma” and “fish tank granuloma.” Infections are normally localized to the skin and occur following trauma, with the limbs and extremities most often infected. Nodules or ulcers around the site of inoculation are most common—these are typically single nodules or ulcers but may spread along the lymphatic vessels to regional lymph nodes, similar to sporotrichosis. Rare manifestations of M. marinum include osteomyelitis and disseminated disease, with non-cutaneous manifestations occurring predominately in immunocompromised populations. M. marinum is a slowly progressing disease and the patient may not present to their provider until months after the initial trauma, often complicating the link to water or fish exposures. M. marinum, while it can be transmitted from fish to human, is not transmitted person-to-person and thus does not pose an infection control risk.

AFB cultures are typically incubated at 37 degrees C. However, M. marinum grows optimally at 30C with scant or no growth at 37degrees C. Thus, when skin and soft tissue samples are received for AFB culture, specimens should be held at both 37 degrees C and 30 degrees C to support growth of most NTM and to facilitate the growth of M. marinum, respectively. Recovery of M. marinum in culture from primary specimen takes several weeks to grow, but subculture may take less than 7 days.

M. marinum can be identified by phenotypic characterization, molecular identification, or mass spectrometry. M. marinum is a photochromogen, meaning that colonies possess a yellow pigment when grown in light and no pigment when grown in the dark. This is characteristic of group I of Runyon’s classification. Matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF), which has been used for years in most clinical microbiology laboratories for the identification of bacteria and yeast, can be used to identify Mycobacteria to the complex or species level. While the processing of Mycobacteria for MALDI-TOF is more laborious than for bacterial identification, the use of MALDI-TOF for identification of Mycobacteria is increasing in use. Whole genome sequencing can be performed on pure isolates for molecular identification, although this is more expensive and less common than MALDI-TOF or phenotypic identification.

A thorough history identifying risk factors for M. marinum should be conducted by the physician. Given the length of time for growth, clinical suspicion due to water or fish exposures can guide treatment before the organism grows in culture.  In addition to providing a diagnosis and guiding clinical care, a history of M. marinum infection is also critical to note for latent tuberculosis screening as M. marinum can cross react with tuberculin skin testing and interferon gamma release assays.

References

  1. Aubry A, Mougari F, Reibel F, Cambau E. Mycobacterium marinum. Microbiol Spectr. 2017 Apr;5(2). doi: 10.1128/microbiolspec.TNMI7-0038-2016. PMID: 28387180.
  2. Wu TS, Chiu CH, Yang CH, Leu HS, Huang CT, Chen YC, Wu TL, Chang PY, Su LH, Kuo AJ, Chia JH, Lu CC, Lai HC. Fish tank granuloma caused by Mycobacterium marinum. PLoS One. 2012;7(7):e41296. doi: 10.1371/journal.pone.0041296. Epub 2012 Jul 20. PMID: 22911774; PMCID: PMC3401166.
  3. Franco-Paredes C, Marcos LA, Henao-Martínez AF, Rodríguez-Morales AJ, Villamil-Gómez WE, Gotuzzo E, Bonifaz A. Cutaneous Mycobacterial Infections. Clin Microbiol Rev. 2018 Nov 14;32(1):e00069-18. doi: 10.1128/CMR.00069-18. PMID: 30429139; PMCID: PMC6302357.

Paige M.K. Larkin, PhD, D(ABMM), M(ASCP)CM is the Director of Molecular Microbiology and Associate Director of Clinical Microbiology at NorthShore University HealthSystem in Evanston, IL. Her interests include mycology, mycobacteriology, point-of-care testing, and molecular diagnostics, especially next generation sequencing.

Microbiology Case Study: A 30 Year Old with Worsening Right Middle Finger Pain

Case History

A thirty year old male on dialysis with past medical history of nephrotic syndrome and end-stage renal disease presented to the emergency department due to complaints of severe abdominal pain. Culture of paracentesis fluid confirmed a diagnosis of bacterial peritonitis, and he was admitted for intra-peritoneal antibiotic therapy. During his admission however, he also complained of progressive worsening right middle finger pain and swelling after smashing his finger in a door one month prior. Upon exam the finger appeared visibly swollen with slight discoloration, but exhibited full range of motion (Image 1A). Imaging revealed osteomyelitis of the intermediate and distal phalanx, including involvement of the distal interphalangeal joint (Image 1B). Additionally, a small abscess was detected (Image 1B, inset) requiring incision and drainage (I&D). Following the I&D, the patient was discharged home on broad spectrum antimicrobial therapy pending results of intra-operative cultures. The patient was readmitted the following week for severe electrolyte imbalance due to a missed dialysis session. His right middle finger continued to cause him significant pain, and the I&D site appeared poorly healing with excess fluid drainage (Image 1C).

Image 1. A: Right distal right middle finger swelling at admission. B: MRI with IV contrast of the right hand revealed osteomyelitis with small abscess formation (inset). C: Right middle figure I&D site at readmission.

Laboratory Identification

Bacterial, fungal and mycobacterial cultures of purulent material from the I&D were obtained. Following four days of incubation, creamy white yeast-like colonies appeared on blood and chocolate agar plates at 35°C (Images 2A, B). Fungal medias grew colonies which were initially white (Image 2C), but became more wrinkled with increased central dark coloration and white periphery (Image 2D) within two weeks at 30°C. Cellophane tape preparations with lactophenol cotton blue were performed for microscopic evaluation of the mature colonies from potato dextrose agar. Narrow, septate hyphae with slender, tapering conidiophores were observed (Image 3A). The conidiophores had many small, tear-shaped hyaline conidia forming “daisy-like” clusters (Image 3B). Based upon these morphologic characteristics, the organism was identified as member of the Sporothrix schenckii complex.

Image 2. Culture of purulent material obtained from right middle finger I&D. A,B: Creamy white yeast-like colonies on blood and chocolate agar plates incubated at 35°C. Small white colonies (C) matured into moist, wrinkled colonies with central dark coloration (D) on fungal medias after 2 weeks’ incubation at 28-30°C.
Image 3. Lactophenol cotton blue (LCB) prep of fungal growth from plate in Figure 2D. Thin, septate hyphae were observed with tapered conidiophores and characteristic “daisy-like” clusters of conidia.

Discussion

Sporotrichosis (also known as rose gardener’s disease) is caused by dimorphic molds belonging to the Sporothrix schenckii complex. These fungi inhabit soil, plant matter and decaying vegetation, thus explaining their traditional association with gardening. As thermally dimorphic fungi, members of the Sporothrix schneckii complex exhibit both yeast and filamentous mold forms. The yeast colonies are smooth with cream or tan coloration when grown close to body temperature (35℃). Microscopically, the yeasts appear as round, oval and fusiform budding cells3,4 best visualized with GMS or PAS stains due to their relative infrequency when observed in tissue. These are commonly referred to as “cigar bodies”, and are typically seen on histopathology, sometimes with accompanying pyogranulomatous inflammation or suppurative granulomas.2 By contrast, the mold colonies begin small, white to pale-orange and become moist, wrinkled, leathery, or velvety and often darken to a salt-and-pepper brown-black as the mold ages. Microscopically, narrow, septate hyphae with slender, tapering conidiophores can be visualized. The conidiophore apex is often slightly swollen and bears many small, tear-shaped or almost round hyaline conidia (2-3 X 2-6 µm) on delicate hair-like denticles, forming the characteristic “rosette” or “daisy-like” clusters in young cultures.1,3,4 This also illustrates the etymology of the organism: from latin, spor (spore) + thrix (hair).5

The most common presentation of sporotrichosis is a lymphocutaneous form originating from a puncture or trauma that inoculates contaminated material into the wound. A pustule forms in subcutaneous tissue followed by ulceration and development of secondary lesions via lymphatic spread in a distal to proximal fashion.2 This form often occurs in immunocompetent individuals. More uncommon forms include osteoarticular, pulmonary, and disseminated infections, usually associated with immunocompromised hosts and alcoholics. In our case of osteoarticular sporotrichosis, the patient suffered from nephrotic syndrome which is often associated with immunodeficiency. The key inciting event to the development of his infection was likely the traumatic injury to his finger which was followed by wound inoculation of soil/material containing the organism. Following his diagnosis, the patient revealed that he was an avid gardener.

In conclusion, the patient was started on oral itraconazole initially for medical management. However, due to continued pain after a few weeks of therapy, the patient opted for amputation of the involved portion of the finger. Following successful amputation, the patient is to remain on itraconazole for one year due to concern for possible surgical margin involvement. He has healed well to date, with no continued pain in the remaining portion of the finger.

References

  1. Love GL, Ribes JA. 2018. Color Atlas of Mycology, An Illustrated Field Guide Based on Proficiency Testing. College of American Pathologists (CAP), p. 214-218.
  2. Pritt BS. 2018. Atlas of Fundamental Infectious Diseases Histopathology. College of American Pathologists (CAP), p. 120-122.
  3. Walsh TJ, Hayden RT, Larone DH. 2018. Larone’s Medically Important Fungi, A Guide to Identification. 6th Ed. ASM Press, p. 53, 178-180.
  4. St-Germain G, Summerbell R. 2011. Identifying Fungi: A Clinical Laboratory Handbook, 2nd Edition, Star Publishing Company Inc., pg. 238-239.
  5. Sellera, FP and Larsson, CE. 2019. Sporothrix schneckii. Etymologica. Emerg. Infect. Dis. 25(9):1631.

John Markantonis, DO is the current Medical Microbiology fellow at UT Southwestern and will be completing his Clinical Pathology residency in 2022. He is also interested in Transfusion Medicine and parasitic diseases.

Samira Peyrovi, M(ASCP)CM is a medical technologist at UT Southwestern Medical Center. She graduated with a degree in biology from Texas Women’s University and has worked in the microbiology section at Clements University Hospital for the past 5 years.

-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.

Microbiology Case Study: A 59 Year Old with History of Rheumatoid Arthritis

A 59 year old male with a history of rheumatoid arthritis, hepatitis C, non-small cell lung carcinoma presents to his rheumatologist. His joint pain has responded well to adalimumab and methotrexate, but over the past month his left elbow has become increasingly painful, despite multiple corticosteroid injections there. He does not report any fevers or chills. On physical exam, his left elbow is warm and swollen. Synovial fluid is aspirated and sent for gram stain and culture.

Image 1. Initial Gram stain findings.
Image 2. Growth on potato flake agar.

The yeast was identified as Candida parapsilosis, a common environmental species of Candida that is becoming increasingly prevalent as a cause of invasive candidal disease. It is easily transmitted by contact and is a source of nosocomial infections. It has the ability to form a biofilm and, thus, has a predilection for indwelling catheters and prosthetic devices. Highest-risk groups include immunocompromised patients, surgical patients, and very low-birth weight neonates.

The microbiology findings exemplify how fungi can be broadly categorized based on how they grow (as a yeast or a mold) in the host and in culture (a cooler environment). In this particular case, the initial gram stain (figure 1), representative of the phenotype in the host, demonstrates neutrophils with intracellular organisms with visible cell walls, suggestive of an active yeast infection with ongoing phagocytosis. The potato flake agar (figure 2) is also growing a yeast. Thus, the pathogen is classified as a yeast. In contrast, molds are characterized by the development of hyphae, which give them their classic “fluffy” appearance in culture (absent in this case). A clinically important subset of this group, the dimorphic fungi, are defined by a yeast phenotype in the host and a mold phenotype in culture.

References

  1. McGinnis, M, and S Tyring. (1996). Medical Microbiology (4th Edition). Galveston, TX: University of Texas Medical Branch at Galveston.
  2. McPherson, R, and M Pincus. (2011). Henry’s Clinical Diagnosis and Management By Laboratory Methods (22nd Edition, pp. 1155-1184). Philadelphia, PA: Elsevier Saunders.
  3. Trofa, D, A Gacser, and J Nosanchuck. “Candida parapsilosis, an Emerging Fungal Pathogen.” Clin Microbiol Rev. 2008 Oct; 21(4): 606-625.

-Frederick Eyerer, MD is a 3rd 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.

Microbiology Case Study: An 83 Year Old Man with History of Dementia

An 83 year old man presents to an emergency department after an unwitnessed fall and being found down for an undetermined amount of time.  His past medical history includes dementia and type II diabetes mellitus.  A tick is found in his left groin.  Workup reveals mild rhabdomyolysis, mild transaminitis, thrombocytopenia, and multiple infiltrates on chest x-ray, concerning for aspiration pneumonia.  His hospital course is complicated by fever, hypotension, and atrial fibrillation with rapid ventricular response.  Representative images from his peripheral blood smear are shown below.

PCR confirmed the diagnosis of Anaplasma phagocytophilum.  The patient recovered after a 10-day course of doxycycline.

Differentiating Tick-Borne Intracellular Microorganisms

The clinical findings of recent tick exposure, thrombocytopenia, transaminitis, and acute illness are consistent with acute infection with either Ehrlichia or anaplasma.  The images depict morulae (microcolonies of bacteria) within the cytoplasm of neutrophils.  A. phagocytophilum, Ehrlichia chaffeensis, and Babesia microti can all be seen intracellularly on peripheral blood smears.  A. phagocytophilum preferentially infects neutrophils, while E. chaffeensis infects monocytes and B. microti infects erythrocytes.  Unlike A. phagocytophilum and E. chaffeensis, which both appear as intracellular morulae (clusters of bacteria), B. microti is a protozoa whose trophozoite forms are seen in red blood cells as delicate rings or grouped in tetrads (the characteristic “Maltese cross”).

The finding of morulae in granulocytes has a sensitivity of 20-80% for anaplasmosis within the first week of infection.  However, morulae within granulocytes are also found in infection with Ehrlichia ewingii (human granulocytotropic Ehrlichiosis, distinct from human monocytotropic Ehrlichiosis caused by E. chaffeensis), a less common relative of E. chaffeensis that is also transmitted by the Lone Star tick (Amblyomma americanum) and appears to preferentially infect immunocompromised hosts.  For E. chaffeensis, the sensitivity of finding morulae in monocytes is only 7-17% amongst immunocompetent hosts.

References

  1. Chapman, AS,  JS Bakken, SM Folk, et al.  “Diagnosis and management of tickborne rickettsial diseases: Rocky Mountain spotted fever, ehrlichioses, and anaplasmosis–United States: a practical guide for physicians and other health-care and public health professionals.” MMWR Recomm. Rep. 2006; 55(RR-4):1.
  2. McPherson, R, and M Pincus.  (2011).  Henry’s Clinical Diagnosis and Management By Laboratory Methods (22nd Edition, pp. 1073-1074, 1205).  Philadelphia, PA:  Elsevier Saunders.
  3. Paddock, CD, SM Folk, GM Shore, et al.  “Infections with Ehrlichia chaffeensis and Ehrlichia ewingii in persons coinfected with human immunodeficiency virus.” Clin Infect Dis. 2001; 33(9):1586-94.
  4. Schotthoefer, AM, JK Meece, LC Ivacic, et al.  “Comparison of a Real-Time PCR Method with Serology and Blood Smear Analysis for Diagnosis of Human Anaplasmosis: Importance of Infection Time Course for Optimal Test Utilization.” J Clin Microbiol. 2013 Jul; 51(7): 2147-2153.

-Frederick Eyerer, MD is a 3rd 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.

Microbiology Case Study: Middle Aged Man with Altered Mental Status

Case History

A middle aged male presented to the emergency department with a several day history of altered mental status, insomnia, and lethargy. His family stated that he also had possibly had a seizure. Upon arrival to the emergency department he was febrile to 102°F, hypoxic, but denied shortness of breath, cough, nausea, and vomiting. While in the care of the emergency department he had several witnessed seizures requiring Ativan treatment, so the decision was made to admit the patient for 24-hour EEG monitoring. Past medical history was unremarkable. The patient is a smoker. His hobbies include working as a mechanic and working outdoors in his garden.

Initial brain imaging on MRI demonstrated cortical thickening/gyral swelling with associated signal abnormality within bilateral mesial temporal lobes that is seen with mild associated diffusion abnormality. A lumbar puncture was ordered and his CSF analysis demonstrated normal glucose (73 mg/dL [reference range: 40-80 mg/dL]), normal protein (32 mg/dL [reference range: 15-45 mg/dL]), and normal nucleated cells [reference range: 0-8/mm3]. Bacterial, fungal, and AFB culture as well as PCR for herpes simplex virus and varicella zoster virus were ordered. The CSF bacterial, fungal, and AFB cultures showed no growth of any microorganisms, and PCR results were negative for HSV and VZV. What additional infectious etiologies would you like to test for?

Discussion

Serology studies were ordered which demonstrated positive IgM and IgG antibodies for West Nile virus.

West Nile virus is a member of the Flavivirus genus. It is spread through mosquito bites, and birds are the primary reservoir for this virus. The incubation period for West Nile virus is 4-10 days. Typically, about 80% of those infected with the virus will be asymptomatic. In 20% of cases, the patient will develop a febrile illness, with possible symptoms of headaches, body aches, weakness, joint pain, and fatigue. About 1 in 150 of those infected will develop illness involving the central nervous system. In these cases, symptoms can include high fever, headache, neck stiffness, confusion, seizures, and coma. Death occurs in 10% of those with involvement of the central nervous system. The most important risk factor for death is age with patients over 70 years of age being most at risk.

Diagnosis of West Nile virus generally made by detection of IgM and IgG in the serum and/or CSF. IgM can be detected 3-8 days post symptom onset and remains positive for 2-3 months in the CSF, and occasionally longer in the serum. Diagnosis is made by detection of IgM antibodies or conversion of IgG antibodies, while detection of IgG antibodies in isolation indicates a prior infection. False-positive results can occur in CSF specimens contaminated with blood that is IgM antibody positive. Patients who have been infected with or vaccinated against other flaviviruses can have false-positive serum antibodies. Plaque reduction neutralization test (PRNT) assay can be performed to rule out cross-reactivity with other flaviviruses. RT-PCR testing on CSF is also available for detection of West Nile virus RNA. The sensitivity of CSF RT-PCR is low in immunocompetent patients, but increased in immunocompromised patients due to prolonged viremia.

West Nile virus cases could increase in the future as a consequence of climate change. Currently, most cases of West Nile virus occur between June and September and cases have been reported in all 48 lower states. As climate change progresses, average temperatures are expected to increase around the world. As temperature increases, summers become longer, which means a longer season for mosquitoes. Laboratory studies also demonstrate that the virus replicates faster in warmer temperatures. Precipitation patterns are also expected to change in the future. The effect of precipitation is not as clear as that of temperature, but it can still play a role. In areas with more precipitation, increasing amounts of water could provide new breeding grounds for mosquitoes. On the other hand, extra rain can dilute some of the nutrients that the mosquitoes need. In areas with less precipitation, drought conditions can cause rivers to dry up, creating the pools of standing water that give rise to mosquitoes. In addition, as bodies of water become smaller, birds and mosquitoes will be in closer proximity, facilitating faster spread of the virus. We must be prepared for the possibility that West Nile virus cases may be on the rise in the future.

The patient continued to suffer from seizures and required continuous sedation with phenobarbital. For seizure prophylaxis he was given a combination of Keppra and Vimpat, with the dosage adjusted as needed. Throughout his entire hospital course he was monitored on EEG. For the first week of his stay, his EEG results demonstrated seizure like activity, requiring continued use of anti-seizure medication and sedation. After about one week, his seizure activity began to show improvement, and the process of decreasing his medication had begun.

After several days of improvement, his recover was complicated by abdominal compartment syndrome requiring laparotomy. After this, the patient’s seizure activity began to worsen. Within a few days, the patient became hypotensive, and broad spectrum antibiotics were given to protect against infection. Cultures were taken but no infection was identified. The patient ultimately developed shock and passed away.

References

  1. Paz, S. (2015). Climate change impacts on West Nile virus transmission in a global context. The Royal Society. doi:https://royalsocietypublishing.org/doi/10.1098/rstb.2013.0561
  2. Murray, P. R., Rosenthal, K. S., & Pfaller, M. A. (2021). Togaviruses and Flaviviruses. In Medical microbiology. Edinburgh: Elsevier.
  3. Miller, J. Michael, et al. “A guide to utilization of the microbiology laboratory for diagnosis of infectious diseases: 2018 update by the Infectious Diseases Society of America and the American Society for Microbiology.” Clinical Infectious Diseases 67.6 (2018): e1-e94.

-Robert Toelke, MD is a 1st year clinical and anatomic pathology resident at University of Chicago (NorthShore).

-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: A 41 Year Old Male with Pneumonia

Case Description

A 41 year old male with a past medical history significant for HIV presented to the emergency department with complaints of a fever, shortness of breath, cough, myalgias, diarrhea, and dark urine for five days. Upon presentation, he was found to meet sepsis criteria for fever of 105° F, white blood cell count of 19 x 109 /L, and tachycardia. Physical exam was unrevealing. Chest x-ray revealed a lobar pneumonia and a CT chest showed ground-glass opacities with superimposed interlobular thickening and intralobular septal thickening, commonly referred to as “crazy-paving” (Image 1). Initial laboratory assessments included CBC, CMP, blood and sputum cultures, and a T-spot. A legionella urine antigen test was ordered a short time later following an infectious disease consult.

CBC and CMP were significant for leukocytosis (19 x 109 /L), hyponatremia (130 mmol/L), and transaminitis (AST: 97, ALT: 91). Blood and sputum cultures were negative as was the T-spot. Ultimately, the diagnosis of Legionnaire’s Disease was made by the positive urine antigen test.

Image 1. Computed tomography images of the chest without contrast demonstrates consolidation and “crazy paving” patterns of radiographic pathology.
Image 2. Representative example of Legionella pneumophila on Buffered Charcoal Yeast Extract (BCYE) media with small wet gray colonies.

Discussion

Legionella is a genus of aerobic, gram negative, intracellular pathogens that are most often found in soil and water.1 There are over 60 known species of Legionella with each consisting of a varied number of serogroups. At least 26 of these species are pathogenic in humans, however only a few are responsible for the vast majority of known cases.2 In North America, upwards of 90% of cases are caused by L. pneumophila, and more specifically its serogroup 1. In Australia and New Zealand, L. longbeachae is the predominant human pathogen.3 Legionella infections most commonly cause community-acquired pneumonia after inhalation of aerosols and can less frequently cause a self-limited febrile syndrome known as Pontiac Fever. Characteristic signs that may cause a treating physician to think of Legionella infection include a constellation of symptoms that include diarrhea, hyponatremia, and elevated liver function tests.4

This case reviews a typical presentation of Legionnaire’s disease and highlights several diagnostic pearls worth remembering. Despite commonly being thought of as an exotic pathogen, Legionella is known to cause between 2%-15% of community-acquired pneumonia cases in the United States and Europe.5 The gold standard for diagnosis is culture of lower respiratory secretions, however it is a fastidious organism that is not easily picked up on gram stain or grown on standard media. When culture is attempted, nutrient enriched BCYE agar is required and the timeframe for growth must be increased to 5 to 7 days for L. pneumophila and 14 days for non-pneumophila strains (Image 2).6 Adding to the difficulty in detection, Legionella is easily treated by empiric therapies, such as macrolides, that cover atypical infections; therefore, delays in testing further reduce sensitivity. The urine antigen test does help to overcome this problem as it can detect infection within 2-3 days of symptom onset and remains positive for at least 1 month following resolution of the illness.6

References

  1. Edelstein PH and Roy CR. Legionnaires’ Disease and Pontiac Fever. In: Mandell, Douglas, and Bennett’s Principles and Practice of Infectious Diseases, 8, Bennet JE, Dolin R and Blaser MJ (Eds), Elsevier, Pennsylvania 2015.
  2. National Center for Biotechnology Information Taxonomy Browser. https://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Tree&id=444&lvl=3&keep=1&srchmode=1&unlock (Accessed on April 14, 2021).
  3. Yu VL, Plouffe JF, Pastoris MC, et al. Distribution of Legionella species and serogroups isolated by culture in patients with sporadic community-acquired legionellosis: an international collaborative survey. J Infect Dis 2002; 186:127.
  4. Robert R. Muder, L. Yu Victor, Infection Due to Legionella Species Other Than L. pneumophilaClinical Infectious Diseases. 2002; 35(1):990-998. doi.org/10.1086/342884
  5. Chahin A, Opal SM. Severe pneumonia caused by Legionella pneumophila: differential diagnosis and therapeutic considerations. Infect Dis Clin North Am. 2017;31(1):111-121. doi:10.1016/j.idc.2016.10.009
  6. Mercante JW, Winchell JM. Current and emerging Legionella diagnostics for laboratory and outbreak investigations. Clin Microbiol Rev. 2015;28(1):95-133. doi:10.1128/CMR.00029-14

-Allen Green is a first year Clinical Pathology resident at UT Southwestern. He has broad interest in laboratory medicine and Transfusion medicine.

-Dominick Cavuoti is a full professor at UT Southwestern and practices both Medical Microbiology, Infectious Disease Pathology and Cytology.

-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.

Microbiology Case Study: A Patient, A Toilet, and a Worm

Case History

A patient presents with a worm they found in the toilet.

Image 1. The offender.
Image 2. Mouth parts of the worm.

Discussion

This is Ascaris lumbricoides, a roundworm. Distinctive morphologic features include tapered ends, mouthparts consisting of three prominent lips (pictured in image 2), and a length of up to 35cm for females. The adults live in the duodenum and proximal jejunum. The eggs have an irregular external mamillated outer shell that gives them a roughened outer surface. Clinically, infection can range from asymptomatic to severe disease, in which the larvae, hatched from ingested eggs, migrate from the small bowel through the circulatory system to the lungs, where they mature in the alveolar capillary bed and cause Ascariasis pneumonitis (Löffler syndrome).

Other diagnostic considerations include Enterobius vermicularis (pinworm), Lumbricus terrestris (earthworm), Trichuris trichiura (whipworm), and the hook worms, Necator americanus and Ancylostoma duodenale.

Enterobius vermicularis, the pinworm, is the most common helminth infection in the United States. Clinically, the classic presentation is a child with pruritus ani. Females measure up to 1.3 cm in length and have a pointed posterior end, and both sexes have lateral alae and a prominent esophageal bulb. The worm in this case is far too large to be a pinworm.

The earthworm, Lumbricus terrestris, is soil-dwelling and non-pathogenic but occasionally encountered in the laboratory for identification purposes. Key morphologic features include a segmented body with no distinctive mouthparts and a clitellum (a mating organ that is a non-segmented portion of the body and often a different color from the rest of the body).

Trichuris trichiura, the whipworm, have a classic whip-like appearance with long, narrow anterior ends that anchor the worm to the large intestine, where they can remain for up to 10 years. Both males and females measure up to 5.0 cm in length, and diagnosis is often made by identification of the eggs, which are football-shaped and have polar plugs at both ends. Clinically, trichuriasis can cause dysentery-type symptoms and, in heavily infected children, can lead to rectal prolapse.

Necator americanus and Ancylostoma duodenale are the hookworms. Adult females measure up to 1.2 cm, and these two species are differentiated by examination of the mouthparts: Necator americanus has cutting plates, while Ancylostoma duodenale has cutting teeth. In addition to the large size difference between hookworms and roundworms, the lamprey-like appearance of these mouthparts is notably different from the “fleshy lips” of Ascaris. Hookworms and roundworms, however, are similar in that their larvae have the ability to migrate through tissue to the blood stream then the lungs, where they can cause Löffler syndrome and are expectorated then swallowed before reaching the small bowel. Unlike Ascaris, the larvae of which hatch from ingested eggs and penetrate the host through the bowel wall to get to the lung capillary beds where they can mature, hookworm larvae hatch outside the body and, on contact with a host (once again, lamprey-style), directly penetrate the skin, enter the circulation, travel to the lungs, then migrate up the bronchial tree to be swallowed. If ingested, Ancylostoma larvae can mature into adults in bowel without needing to migrate through the lungs.

References

  1. Centers for Disease Control and Prevention. “Stool Specimens – Intestinal Parasites: Comparative Morphology Tables.” https://www.cdc.gov/dpdx/diagnosticProcedures/stool/morphcomp.html. Last reviewed May 3, 2016. Accessed April 2, 2021.
  2. “Earthworms.” University of Pennsylvania. https://www.sas.upenn.edu/~rlenet/Earthworms.html. Accessed April 2, 2021.
  3. McPherson, R, and M Pincus. (2011). Henry’s Clinical Diagnosis and Management By Laboratory Methods (22nd Edition, pp. 1218-1220). Philadelphia, PA: Elsevier Saunders.

-Frederick Eyerer, MD is a 3rd 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.

Microbiology Case Study: A 55 Year Old Man with New Onset Neurologic Deficits

Clinical History

A 55 year old male with a 43-pack-year smoking history was transferred to our hospital for evaluation of new onset neurologic deficits including slurred speech, aphasia, and right upper extremity diminished dexterity and neglect. CT chest was remarkable for a mass in the superior segment of the left lower lobe. Needle core biopsy of the lung mass revealed poorly differentiated non-small cell carcinoma. Head MRI demonstrated an enhancing mass in the left frontoparietal junction that was concerning for metastasis from a lung primary. The patient was started on chemotherapy as an outpatient. Follow-up imaging showed growth of the brain mass. A biopsy of the brain mass showed no evidence of metastasis, only “reactive brain with foci of dense mixed inflammation and filamentous bacteria consistent with abscess.”

Image 1. Head MRI demonstrating left frontoparietal mass.

Laboratory Findings

A portion of the brain biopsy was submitted for bacterial smear and culture. The aerobic culture grew chalky white colonies that, when stained with modified acid-fast stain, showed modified acid-fast positive filamentous bacteria, suspicious for Nocardia spp. Bacteria of similar morphology were also seen in the surgical pathology specimen when stained for modified AFB and with GMS.

Image 2. Blood agar plate growing chalky white colonies.
Image 3. Modified acid-fast positive filamentous bacteria at 1000x.

Disscussion

Nocardia is a genus of aerobic, catalase positive, saprophytic bacteria often found in the environment, but that can also be considered as normal flora on skin and in the respiratory tract. Nocardia species are variably acid-fast; for proper identification they must be stained with a modified acid-fast procedure (Fite, Kinyuon), using a weaker decolorizing acid. Nocardia will be negative by traditional acid-fast staining procedures (Ziehl–Neelsen). When Gram stained, Nocardia will appear as branching filamentous gram-positive bacilli with a “beaded” staining pattern (as if a string of beads). 

Multiple species are considered human pathogens, including N. asteroides, N. brasiliensis, N. cyriacigeorgica, N. farcinica, and N. nova. These organisms can cause disease in immunocompromised patients if inhaled or inoculated via trauma. If there is an established pulmonary infection, Nocardia may spread hematogenously, often infecting the brain. 

Central nervous system nocardiosis may occur in any region in the brain and can present with mass effect symptoms without typical infectious symptoms, as in our patient. Prognosis varies based on the extent of disease and the cause of a patient’s immunosuppression. Treatment of CNS nocardiosis usually begins with an induction phase of intravenous TMP-SMX and imipenem for 3-6 weeks or until there is clinical improvement. Once the patient improves, they can be switched to oral therapy with a sulfonamide and/or minocycline and/or amoxicillin-clavulanate.  

References

  • Beaman BL. Nocardia Species : Host-Parasite Relationships. 1994;7(2):213-264.
  • Spelman D. Clinical manifestations and diagnosis of nocardiosis. In: Sexton DJ, Mitty J, eds. UpToDate. UpToDate, Inc.
  • Spelman D. Microbiology, epidemiology and pathogenesis of nocardiosis. In: Sexton DJ, Mitty J, eds. UpToDate. UpToDate, Inc.
  • Spelman D. Treatment of Nocardiosis. In: Sexton DJ, Mitty J, eds. UpToDate. UpToDate, Inc.
  • Tille, Patricia M., PhD, BS, MT(ASCP) Facs. Bailey & Scott’s Diagnostic Microbiology. 14th ed. Elsevier; 2017.

-Michael Madrid, 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.

Microbiology Case Study: A 53 Year Old Man with Confusion, Auditory Hallucinations, and Hearing Loss

Clinical History

A 53 year old male with a past medical history significant for dermatomyositis, antisynthetase syndrome, and atrial fibrillation with rapid ventricular response presented with a chief complaint of worsening confusion, auditory hallucinations, and hearing loss. Pertinent medications included prednisone and mycophenolate. Head MRI demonstrated leptomeningeal enhancement and hydrocephalus. A lumbar puncture was performed, with CSF results as follows:

Laboratory Findings

CSF was sent to the microbiology lab for bacterial and fungal smears and cultures. No organisms were seen on the Gram stain.

Within 3 days, however, rare colonies of yeast were growing on both the bacterial and fungal media. The yeast was identified as Cryptococcus neoformans using the in-house MALDI-TOF mass spectrometry instrument.

Image 1. Budding C. neoformans at 1000x.
Image 2. Colonies of C. neoformans on potato flake agar.

Discussion

Cryptococcus neoformans is an environmental saprophytic yeast that can be found around the world, although it is often associated with avian droppings.1 The cell is surrounded by a polysaccharide capsule that protects it from environmental hazards and, once within the host, from phagocytosis.2 Additionally, the cell wall of C. neoformans contains melanin due to the presence of the phenol oxidase enzyme, which assists in the formation of melanin from various phenolic substrates.1 Both the polysaccharide capsule and the melanin-containing cell wall can be helpful in the laboratory identification of C. neoformans.

If inhaled, Cryptococcus neoformans can cause disease (cryptococcosis) in immunocompromised patients. The most significant risk factor is AIDS, however any cause of immunodeficiency can be a risk factor, including long-term steroid therapy, organ transplantation, malignancy, and liver disease.1 Once inhaled, the organism spreads hematogenously and tends to favor the central nervous system, causing cryptococcal meningoencephalitis.1

Prognosis for patients with cryptococcosis can vary widely. In AIDS-associated CNS cryptococcosis, predictors of mortality include abnormal mental status, cerebrospinal fluid antigen titer >1:1024 by latex agglutination or >1:4000 by lateral flow assay, and CSF white blood cell count <20/µL.1 The prognosis for patients who are immunocompromised for other reasons depends on the cause of their immunosuppression.1

Treatment of patients with cryptococcal meningoencephalitis consists of an induction phase with amphotericin B and flucytosine followed by a consolidation phase with fluconazole then a long-term maintenance phase with a smaller dose of fluconazole.3

References

  1. Jobson M. Microbiology and epidemiology of Cryptococcus neoformans infection. In: Post T, ed. UpToDate. UpToDate, Inc. Accessed March 13, 2021. https://www.uptodate.com
  2. Tille, Patricia M., PhD, BS, MT(ASCP) Facs. Bailey & Scott’s Diagnostic Microbiology. 14th ed. Elsevier; 2017.
  3. Perfect JR, Dismukes WE, Dromer F, et al. Clinical practice guidelines for the management of cryptococcal disease: 2010 update by the infectious diseases society of America. Clin Infect Dis. 2010;50(3):291-322. doi:10.1086/649858

-Michael Madrid, 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.