Microbiology Case Study: A 58 Year Old Female with Fever, Headache, and Vomiting

Case History

A 58 year old female presented to the emergency department with complaints of a fever (reaching 102.9°F) and headache with associated nausea and vomiting for the past 24 hours. Her past medical history was significant for a resection of a recurrent hemangiopericytoma by the neurosurgery service three weeks prior. The patient also noted clear drainage from this surgical site which had begun 5 days ago. Other symptoms noted at presentation included decreased appetite and dehydration. She denied back & neck pain, photophobia or stroke and seizure-like symptoms. Her vital signs were all within normal limits. On physical exam, a healing surgical wound was noted in the posterior auricular area with clear drainage, but no blood or exudates were visualized.  She had no tenderness when her spine was palpated and neurologic exam showed a left sided facial droop and tongue deviation which were noted previously and attributed to her multiple central nervous system surgeries. Complete blood count (CBC) showed a mild increase in white blood cells and anemia. An external ventricular drain was placed and cerebral spinal fluid (CSF) was sent to the microbiology lab for culture. Blood cultures and a swab from the surgical wound were also collected.

Laboratory Identification


Image 1. Gram stain of the cytospin CSF showed many acute inflammatory cells and numerous Gram negative bacilli (1000x).


Image 2. Growth of large, glossy, reddish-orange colonies on sheep blood agar (image taken after 72 hours of incubation).


Image 3. Growth of large, deep red colonies on MacConkey agar (image taken after 72 hours of incubation).


Gram stain of the CSF showed numerous acute inflammatory cells and many Gram negative bacilli (Image 1). Culture of the CSF and wound swab showed large, glossy red colonies on sheep blood and MacConkey agars (Images 2 & 3). Analysis of the colony by matrix assisted light desorption ionization time of flight mass spectrometry (MALDI-TOF MS) identified the organism as Serratia marscescens.


Serratia marscescens is a facultative Gram negative bacillus that is a member of the Enterobacteriaceae family. S. marscesens is ubiquitous in the environment and the most frequent and clinically important species in the genus. Although S. marscesens usually doesn’t cause infection in healthy individuals, it is notorious for colonizing and causing infections in hospitalized patients, particularly those who are immunocompromised, in intensive care units (especially intubated patients) and those with indwelling catheters.  While respiratory infection are most common, S. marscesens has also been implicated in numerous other opportunistic infections such as urinary tract infections, wound infections and septicemia. Brain abscesses and meningitis are less common. S. marscesens has been implicated as the cause of outbreaks in hospitals and can often be traced back to pieces of medical equipment including nebulizers, bronchoscopes, laryngoscopes and contaminated solutions. Person to person transmission is also recognized and thought to be predominantly transmitted via direct contact.

In the laboratory, S. marscesens can be identified by its characteristic non-diffusible red pigment, prodigiosin. Care should be taken when interpreting the lactose reaction on MacConkey agar, as the red pigment may be confused with a positive reaction, while S. marscesens is known to be lactose negative.  As a member of the Enterobacteriaceae family, S. marscesens is able to ferment glucose, reduce nitrate to nitrite and has a negative oxidase reaction. A unique feature of this genus is that all Serratia spp. produce three proteolytic enzymes: lipase, gelatinase, and DNase. Commercial systems, including MALDI-TOF MS, are helpful in the identification of S. marscesens as well.

Treatment of Serratia marscescens infections can be difficult due to various antimicrobial resistance mechanisms, such as expression of extended spectrum beta lactamases (ESBLs), AmpC cephalosporinases and carbapenemases, exhibited by the organism. In the case of our patient, she was empirically started on vancomycin and piperacillin-tazobactam and taken to surgery for wound wash out, removal of hardware and repair of CSF leak. Her antibiotics were changed to meropenem and gentamicin. She was discharged to a rehabilitation facility and received meropenem for a total of 6 weeks.



-David Marbury, MD, is a 3rd year Anatomic and Clinical Pathology resident at the University of Mississippi Medical Center.


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

Microbiology Case Study: An 11 Year Old with Abdominal Pain

Case History

An 11 year-old patient with a history of a relapsed lymphoma presented to the hematology/oncology clinic with worsening abdominal pain. The patient was recently started on metronidazole to treat a C. difficile infection. In the clinic, the patient was found to be hypotensive, hypoxemic and pancytopenic. Blood cultures were drawn and the patient was admitted directly to the pediatric ICU and started on empiric antibiotics.

The blood cultures turned positive with Gram-positive cocci, which went on to produce small, gray, alpha-hemolytic colonies on the blood agar plate (Image 1). The colonies were catalase negative and PYR negative. The isolate was analyzed by a Bruker MALDI-TOF mass spectrometer and was identified as Streptococcus lutetiensis (score 2.19). Susceptibility testing revealed the isolate susceptible to ceftriaxone, penicillin, and vancomycin.


Image 1. 5% sheep blood agar growing small, grey, alpha-hemolytic colonies



S. lutetiensis is part of the complex of organisms previously identified as the Streptococcus bovis group. This group of organisms, which possess the Lancefield Group D antigen, has undergone considerable reclassification schemes as phenotypes and genotypes have been investigated. The original biochemical classification schemes were based on their ability to ferment mannitol as well as the presence or absence of beta-glucuronidase activity. Early observations of the DNA properties from these organisms, such as %-GC base content and DNA-DNA hybridizations, identified six unique clusters of Group D streptococci [1]. One cluster group (cluster group #4) had heterogeneous biochemical phenotypes. A subcluster of this cluster group #4 was separated from the other members of the cluster based upon esculin hydrolysis. This subcluster would later go on to be named S. infantarius, so named as several isolates originated from the feces of human infants [2].

Further DNA-DNA hybridizations and ribotyping analysis led to the declaration of two S. infantarius subspecies: subsp. infantarius and subsp. coli [3]. The 16S rRNA ribotyping was problematic, however, as several species in this genus are 97-99% sequence identical.

In an attempt to address some of the limitations of relying on the 16S rRNA gene, one group analyzed the features of the conserved gene encoding the manganese-dependent superoxide dismutase gene (sodA). They observed substantial differences between S. infantarius subsp. infantarius and S. infantarius subsp. coli [4]. Thus, for the latter organism, a new species of streptococci was proposed: S. lutetiensis. It was named for Lutetia, a historical name for the city of Paris [4].

The species designation S. lutetiensis was not widely accepted, however. Based on further DNA-DNA hybridization experiments and the prior studies of the 16S rRNA, others have rejected the species name “S. lutetiensis” and maintain that it is a subspecies of S. infantarius as previously described [5].

So which name is correct? There appears to be no clear consensus about the designation of these streptococci, whether it is S. infantarius subsp. coli or whether it is another species altogether as S. lutetiensis. The Judicial Commission of the International Committee on Systematic Bacteriology reportedly met to discuss the name changes, however no resolution appears to have been determined [6]. Both names are seen in the literature as well as the names for reference organisms.

The important clinical aspect to recognize is that this organism, as well as the S. bovis group in general, can be a cause of bacteremia, endocarditis, and meningitis in children. Treatment with beta-lactam antibiotics is generally sufficient to cover these organisms.


  1. Farrow, J., et al., Taxonomic Studies on Streptococcus bovis and Streptococcus equinus: Description of Streptococcus alactolyticus sp. nov. and Streptococcus saccharolyticus sp nov. System. Appl. Microbiol, 1984. 5: p. 467-482.
  2. Bouvet, A., et al., Streptococcus infantarius sp. nov. related to Streptococcus bovis and Streptococcus equinus. Adv Exp Med Biol, 1997. 418: p. 393-5.
  3. Schlegel, L., et al., Streptococcus infantarius sp. nov., Streptococcus infantarius subsp. infantarius subsp. nov. and Streptococcus infantarius subsp. coli subsp. nov., isolated from humans and food. Int J Syst Evol Microbiol, 2000. 50 Pt 4: p. 1425-34.
  4. Poyart, C., G. Quesne, and P. Trieu-Cuot, Taxonomic dissection of the Streptococcus bovis group by analysis of manganese-dependent superoxide dismutase gene (sodA) sequences: reclassification of ‘Streptococcus infantarius subsp. coli’ as Streptococcus lutetiensis sp. nov. and of Streptococcus bovis biotype 11.2 as Streptococcus pasteurianus sp. nov. Int J Syst Evol Microbiol, 2002. 52(Pt 4): p. 1247-55.
  5. Schlegel, L., et al., Reappraisal of the taxonomy of the Streptococcus bovis/Streptococcus equinus complex and related species: description of Streptococcus gallolyticus subsp. gallolyticus subsp. nov., S. gallolyticus subsp. macedonicus subsp. nov. and S. gallolyticus subsp. pasteurianus subsp. nov. Int J Syst Evol Microbiol, 2003. 53(Pt 3): p. 631-45.
  6. Beck, M., R. Frodl, and G. Funke, Comprehensive study of strains previously designated Streptococcus bovis consecutively isolated from human blood cultures and emended description of Streptococcus gallolyticus and Streptococcus infantarius subsp. coli. J Clin Microbiol, 2008. 46(9): p. 2966-72.



-I.J. Frame MD, PhD, is a 1st year Clinical Pathology Resident at UT Southwestern Medical Center.

Erin McElvania TeKippe, PhD, D(ABMM), is the Director of Clinical Microbiology at Children’s Medical Center in Dallas Texas and an Assistant Professor of Pathology and Pediatrics at University of Texas Southwestern Medical Center.

Microbiology Case Study: A 28 Year Old Woman with Acute Onset Fever Post Delivery

Case History

A 28 year old woman at 37 weeks and 2 days presented in labor to our ED. After 22 hours, she delivered a healthy baby boy and sustained a second degree perineal laceration requiring repair. On hospital day 2, she reported feeling lightheaded, nauseous and “shaky.” She attempted to walk around the unit but became tremulous and unsteady, requiring assistance to get back into bed. Her vital signs were as follows: febrile at 38.8 C, BP 108/54, HR 104 and normal respiration rate at 12 breaths/min. On exam, she appeared pale and lethargic, and was noted to have a tender uterus on palpation. Based on her presentation and status post SVD, the diagnosis of endometritis was established. Blood cultures were obtained and within 16 hours, blood culture bottles were positive for gram-positive cocci. The patient was started on antibiotic therapy with ampicillin, gentamycin and clindamycin, and clinically improved within 36 hours.

Image 1. Blood culture on blood agar.
Image 2. Blood culture on chocolate agar.


Streptococcus pyogenes is one of the most aggressive pathogens encountered in clinical microbiology. It is a beta hemolytic streptococcus and is notoriously associated with Streptococcal Toxic Shock Syndrome (STSS), necrotizing fasciitis, as well as more benign (yet still problematic) conditions, like Scarlet Fever, Impetigo, Rheumatic heart disease and Acute Post-streptococcal Glomerulonephritis. A gram-positive cocci, it possesses several virulence factors, including protein F, M protein (involved in antigen mimicry leading to valvular heart disease) hemolysins and exotoxins. These factors allow S. pyogenes to attach to and invade epithelial tissue, and in the case of hyalurondiase, potentially use hyaluron as a carbon food source. S. pyogenes agglutinates with Lancefield group A antisera and is pyrrolidonyl arylamidase (PYR) positive and VP, hippurate and CAMP test negative. Penicillin (PCN) remains the drug of choice in treating most S. pyogenes infections. Alternative antibiotic therapy includes macrolides and certain cephalosporins (e.g. cefixime, cefpodoxime). Vancomycin should be used in more severe infections such as sepsis or for patients with a PCN allergy.


-Christina Litsakos is a Pathology Student Fellow at University of Vermont Medical Center.


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


Microbiology Case Study: A 47 Year Old Woman with Three Day History of Fever

Case History

A 47 year old Caucasian female presented to her primary care physician with a three day history of fever (Tmax 102°F), chills and generalized body aches. Her rapid influenza test was negative, but she was treated with oseltamivir for suspected viral infection. Her past medical history was significant for severe mitral regurgitation for which she had had a prosthetic valve replacement two years prior, ischemic cardiomyopathy with recent pacemaker placement one month prior and an undifferentiated connective tissue disease. Her current medications included hydroxychloroquine (Plaquenil) and warfarin. Her symptoms persisted and upon return to clinic, a urinalysis was performed and blood cultures were collected. On physical exam, the pacemaker site was erythematous and tender to palpation. She was started on doxycycline and fluconazole for a presumed urinary tract infection. After 4 days of incubation on the automated instrument, the two aerobic blood cultures bottles were positive and the patient was admitted to the hospital for further workup and therapy.

Laboratory Identification

Microscopic examination from the positive blood culture bottle revealed slender, beaded Gram positive bacilli (Figure 1). No definitive branching was identified. Given the morphology on Gram stain, a Kinyoun stain was performed and revealed red-purple, beaded acid fast bacilli which were consistent with a Mycobacterium spp. (Figure 2). A Mycobacterial Growth Indicator Tube (MGIT), a Lowenstein Jensen slate and blood & chocolate agars were inoculated with specimen. Given that the organism grew after 2 days, a rapidly growing Mycobacterium spp. was suspected (Figure 3). High performance liquid chromatography (HPLC) identified the organism as M. fortuitum.


Figure 1. Gram stain from the positive blood culture bottle showed slender, beaded Gram positive bacilli that were arranged in clumps (100x oil immersion).


Figure 2. Kinyoun stain of the organisms was consistent with acid fast bacilli (100x oil immersion).


Figure 3. Small, off-white colonies grew on chocolate agar after 3 days of incubation at 35°C in a CO2 incubator.


M. fortuitum is a common rapid growing mycobacterial species that is ubiquitous in the environment and tap water. Most common infections due to M. fortuitum include post-traumatic or post-surgical wound infections and it can be associated with the insertion of prosthetic devices including heart valves, artificial joints and rods inserted after fractures. Of the rapid grower group (Runyon Group IV), which includes M. chelonae, M. abscessus and M. mucogenicum, it is M. fortuitum that accounts for approximately 60% of localized cutaneous skin infections and prosthetic device infections most frequently.

In the laboratory, M. fortuitum typically grows after two to five days incubation and appear as small, off-white colonies on a variety of different agars. The organism is typically slender, beaded Gram positive bacilli on Gram stain and positive for acid fast bacilli on a Ziehl-Neelsen or Kinyoun stain. As part of a traditional lab work up, M. fortuitum is arylsulfatase positive and is capable of reducing nitrates. Today a variety of methods, including HPLC, pyrosequencing, sequence analysis and matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS), have become routine identification options. Susceptibility testing of isolates from clinically significant sites should be performed by broth microdilution and includes the following antimicrobials: amikacin, cefoxitin, ciprofloxacin, moxifloxacin, clarithromycin, doxycycline, linezolid, imipenem, meropenem, minocycline, trimethoprim-sulfamethoxazole and tobramycin.

In the case of our patient, it was discovered her pacemaker site was infected and upon further questioning it was discovered she wasn’t able to complete her antibiotic course after device placement due to nausea. A transesophageal echocardiogram showed no evidence of infective endocarditis and she was taken to the operating room for removal of the pacemaker and leads. The site was filled with pus and wound cultures obtained during surgery were consistent with M. fortuitum as well. Repeat blood cultures were negative and she was treated with intravenous amikacin and imipenem as well as oral levofloxacin for an anticipated 6-8 weeks before transitioning to oral therapy.


-Debbie Rigney Walley, MD, is a 1st year Anatomic and Clinical Pathology Resident at the University of Mississippi Medical Center.


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

Microbiology Case Study: A 73 Year Old Man with Back Pain

Case History

A 73 year old man with a history of multiple back surgeries presented with bilateral lower extremity back pain of over greater than one month duration. Prior surgeries included L4/L5 fusion with pedicel screws and a decompression laminectomy one year prior to presentation. Imaging of his spine showed a fluid collection in his lumbar spine and he underwent several tissue biopsies over the course of a month which consistently showed no growth. Despite negative cultures he was treated with doxycycline and levoquin for 30 days. He was transferred to University of Vermont Medical Center (UVMMC) for IR drainage and tissue biopsy of this lumbar abscess as he continued to complain of back pain and had begun to develop bilateral lower extremity weakness. Cultures grow the organism below and close inspection revealed the presence of small feet. The organism was confirmed to be Candida albicans.




Vertebral osteomyelitis due to Candida is rare, however, a review of the literature reveals that most patients have lower thoracic or lumbar spine involvement and over 80% present with >1 month of lower back pain. An elevated white blood cell count is not as sensitive as an elevated erythrocyte sedimentation rate and of all patients, less than a quarter have neurologic signs. Candida albicans was responsible for almost 2/3 of cases and the remaining cases were caused by Candia tropicalis or Candida glabrata.1 Risk factors include IV drug abuse for patients under 25 years old; for elderly patients a central venous catheter, antibiotic use and immunosuppression .1



Miller, D and Mejicano, George. Vertebral Osteomyelitis due to Candida species: Case report and review of the literature. Clinical Infectious Diseases, 2001;33:523-530.

-Agnes Balla, 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 Assistant Professor at the University of Vermont.


Microbiology Case Study: A 14 Year Old Cystic Fibrosis Patient

Case History

14 year old cystic fibrosis (CF) patient was admitted to the hospital for a CF exacerbation with a known rhinovirus infection. The patient reported congestion and cough with production of greenish sputum. The following was isolated from a sputum culture.

Laboratory Culture



Image A. MacConkey Agar plate with mucoid Pseudomonas aeruginosa colonies.


Image B. Kirby-Bauer Method of antimicrobial susceptibility.


Cystic fibrosis patients have complex polymicrobial respiratory flora. Routine cultures may reveal different bacterial species that can contribute to the difficulty in treating and preventing infections in these patients. Staphylococcus aureus, Haemophilus influenza and Pseudomonas aeruginosa are main contributors to the infections found in CF patients.

In this case, we isolated a mucoid Pseudomonas aeruginosa (Image A). Pseudomonas aeruginosa is an oxidase-positive, gram negative rod-shaped bacterium that is commonly found in the environment. It is considered a severe and frequent pathogen in patients with cystic fibrosis. In chronic infections, it is thought that P. aeruginosa can undergo a “mucoid switch” where the bacterium can acquire mutations that lead to the mucoid phenotype. The phenotype is so impressive that excess polysaccharide will often drip onto the lid of the plate when stored upside down during incubation.

One of the key features of mucoid strains of P. aeruginosa is their ability to form biofilms. Biofilms consist of a matrix of polysaccharide, protein and DNA. This provides not only a protective barrier from antibiotics and the immune system, but also may contribute to the growth of other bacteria within the microenvironment.

With all the excess polysaccharide, it can be difficult to standardize the inoculum of mucoid isolates of P. aeruginosa which is an essential starting point for the microbroth dilution method of antimicrobial susceptibility testing. For this reason, susceptibility testing of mucoid isolates is often performed by Kirby-Bauer (KB) disk diffusion method. The KB method is a test of antimicrobial susceptibility that is based on the zone of inhibition surrounding disks that contain antimicrobial drugs (Image B). The strain of mucoid P. aeruginosa isolated in this case was found to be susceptible to aztreonam, ceftazidime, piperacillin/tazobactam and resistant to amikacin, cefepime, ciprofloxacin, gentamicin, levofloxacin, meropenem and tobramycin, by the Kirby Bauer method.

Case Follow Up

The patient was ultimately treated with IV piperacillin/tazobactam, as well as with chest physical therapy and hypertonic saline inhalation. They clinically improved and were ultimately discharged home after a 2 week hospital stay.


The mucoid switch in Pseudomonas aeruginosa represses quorum sensing systems and leads to complex changes to stationary phase virulence factor regulation. Ben Ryall, Marta Carrara, James EA Zlosnik, Volker Behrends, Xiaoyun Lee, Zhen Wong, Kathryn E. Lougheed, Huw D. Williams. PLOS ONE, May 2014, Vol. 9, Iss. 5, Pages 1-11.

Pseudomonas aeruginosa biofilms in cystic fibrosis. Niels Høiby, Oana Ciofu, and Thomas Bjarnsholt. Future Microbiology, November 2010, Vol. 5, No. 11, Pages 1663-1674.

Insights into Cystic Fibrosis Polymicrobial Consortia: The Role of Species Interactions in Biofilm Development, Phenotype, and Response to In-Use Antibiotics. Magalhaes AP, Lopes SP, Pereira MO. Frontiers in Microbiology, January 13, 2017, Vol. 7, Article 2146.

Koneman’s Color Atlas and Textbook of Diagnostic Microbiology. Gary W. Procop, Deirdre L. Church, Geraldine S. Hall, William M. Janda, Elmer W. Koneman, Paul C. Schreckenberger; Gail L. Woods. Seventh Edition. 2017. Pages 343, 1110.



-Megan B. Wachsmann, MD, MSCS, is a 4th year Anatomic and Clinical Pathology Resident and Chief Resident at UT Southwestern Medical Center.

Erin McElvania TeKippe, PhD, D(ABMM), is the Director of Clinical Microbiology at Children’s Medical Center in Dallas Texas and an Assistant Professor of Pathology and Pediatrics at University of Texas Southwestern Medical Center.

Microbiology Case Study: A Bartholin Gland Cyst Culture

Case History

A swab from a bartholin gland cyst was submitted to the microbiology lab with no other clinical history. Culture produced cream-colored, mucoid colonies on blood and chocolate agar after 48 hours of incubation at 35 C (image below). The culture grew a gram negative rod that did not grow on MacConkey agar, a distinguishing characteristic for Weeksella virosa. This ID was confirmed by MALDI-TOF.



Originally described in 1970 as a nonsaccharolytic flavobacterium, Weeksella virosa is an uncommon aerobic gram negative rod isolated most commonly from urine, cervix and vaginal specimens. There have been case reports, however, of isolation from blood and spinal fluid. In rare instances, the bacterium has been associated with pneumonia, bacteremia, peritonitis and UTIs, more often in patients with comorbidities including end stage renal disease, diabetes, and liver disease.

In one study of 100 patients, the organism was isolated in 2% of high vaginal swabs of the female genital tract from both symptomatic and asymptomatic females. In a separate study involving women with a high risk of STDs, the incidence climbed to 15%.

The characteristic yellow tinge of the colonies is secondary to a non-diffusable pigment.  The organism is oxidase positive, indole positive and catalase positive.

There are no species-specific breakpoints. However, CLSI guidelines for other non-enterobacteriacea gram negative rods can be used.  In vitro studies have found the organism to be resistant to aminoglycosides and nitrofurantoin.


Slenker, A et al. Fatal Case of Weeksella virosa Sepsis. Journal of Clinical Microbiology. P4166-4167. Dec. 2012.

-Agnes Balla, 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 Assistant Professor at the University of Vermont.