despite its world - wide omnipresence in soil , a. fumigatus had been considered asexual until studies of an aspergillus - like anamorph isolated from brazilian soil revealed a heterothallic breeding system , and the teleomorph neosartorya fumigata was described .
we describe the first reported case of acute respiratory distress syndrome ( ards ) attributed to neosartorya udagawae .
the mold grew rapidly in culture of both sputum and bronchoalveolar lavage ( bal ) fluid from a previously healthy 43-year - old woman with ards , which developed as the consequence of an acute febrile illness that progressed over 5 days . but despite appearance of fluffy white , non - sporulating mold on a variety of culture media , the colonies did not produce conidiophores , and phenotypic identification was not possible .
the conidia of n. udagawae require longer incubation in the laboratory to germinate , and misidentification of neosartorya sp .
n. udagawae has been emerging as a cause of invasive infections in humans , but our case is the first report of ards associated with neosartorya infection and describes a new clinical entity .
a 43-year - old woman presented to an outlying hospital with complaints of fever and progressive shortness of breath that developed over 5 days .
the patient had a past medical history significant for depression managed with sertraline 50  mg per day , and tobacco use ( cigarettes ) .
she was brought to her local hospital by the emergency medical service after developing profound weakness , with inability to get out of bed .
she was intubated due to respiratory decline and initially required mechanical ventilation with 100% fio2 and positive end - inspiratory pressure ( peep ) of 12  mm / h2o .
she was unable to achieve adequate oxygen saturations with mechanical ventilation and required extra corporeal membrane oxygenation ( ecmo ) at the time of transfer to our facility .
upon arrival at our facility , the patient was hypothermic ( 35.9  c ) .
she was intubated and sedated with a set respiratory rate on mechanical ventilation at 14  breaths / min .
her pulse and blood pressure were 79  beats / min and 86/52  mmhg , respectively ; the fio2 was 99% ; her weight was 93  kg ( 204  lbs ) .
endotracheal and nasogastric ( ng ) tube , foley catheter , and ecmo cannula were in place .
neurologic exam was consistent with medically induced paralysis ; muscle tone , corneal and pupil responses were all normal .
complete blood count was notable for leukocytosis of 17.810  wbc/l ( 78% neutrophils ) , and normocytic anemia ( hgb=10.7  g / dl ) .
the complete metabolic panel was normal , outside of elevated chloride ; hypokalemia ( k=3.1  mg / dl ) and depressed protein and albumin ( 5.4 and 2.6  mg / dl , respectively ) .
an arterial blood gas revealed acidosis and hypoxemia : ph=7.31 ; paco2=43  pao2=45 ; sao2=80% .
semi - upright portable ap chest x - ray was notable for diffuse alveolar opacities .
endotracheal aspirate and three sets of blood cultures and a urine sample were submitted to the microbiology lab for culture .
she was empirically started on oseltamivir 75  mg q 24  h via the ng , along with intravenous azithromycin and ceftriaxone initiated at the outlying hospital . during the patients first 24  h in the icu , she progressed from hypothermia to a febrile state ( 39.1  c ) , resolving leukocytosis ( 10.910  wbc/l ) , and an unchanged physical exam .
hiv elisa , mycoplasma serology , and fungal immunodiffusion antibody testing for aspergillosis , histoplasmosis , blastomycosis , and coccidiomycosis were performed , and all returned negative / non - reactive . a nasopharyngeal swab for multiplex pcr for respiratory viral and intracellular bacterial pathogens ( film array , biofire , inc . )
day two , the patient was noted to have a normal body temperature , but remained on paralytics in a medically induced coma .
one additional blood culture was obtained , and ebv and cmv serologies were performed ; all returned negative .
cultures of an endotracheal tube aspirate ( eta ) that were obtained on admission were notable for growth of a fluffy , sterile white mold with yellow center on standard blood agar media . on hospital day four , endoscopic bronchoscopy was performed .
bal fluid was collected from the left and right lower lobes ( lll & rll ) of the lung and submitted for gram and gomori methenamine silver ( gms ) staining , which were negative .
histoplasma and legionella antigen testing on the bal fluid was negative , but an aspergillus galactomannan antigen was positive ( > 3.75  ng / ml ) .
pcr for influenza ( cepheid genxpert ) as well as multiplex pcr for respiratory pathogens ( film array , biofire , inc . ) were both negative . again , within 48  h , a similar appearing fluffy , white mold grew in cultures of both lll and rll bal fluid samples on all media ( blood agar , sabouraud agar , and mycosel agar ( becton dickinson ) ; a selective medium containing cycloheximide and chloramphenicol to inhibit bacterial growth ) .
potato dextrose agar ( pda ) was inoculated with a sample of mold , but serial lactophenol cotton blue prep exams of the non - sporulating cultures were non - diagnostic , although clearly not consistent with mucormycosis .
empiric treatment with lipid formulation of intravenous amphotericin b was initiated on hospital day 5 due to failure to improve on broad spectrum empiric antibiotic treatment . over the next 48 
h respiratory status was unchanged and oxygenation parameters and radiographic imaging showed no improvement . on hospital day 7 , following two iv doses of liposomal amphotericin b , repeat bronchoscopy was performed .
gram and gms stains , and histoplasma antigen testing were repeated , and returned negative for yeast , pneumocystis or fungal elements .
methylprednisolone 125  mg iv every 8  h was added to the empiric iv amphotericin b for treatment of possible allergic bronchopulmonary aspergillosis ( apba ) .
mold was not cultured from either bal fluid sample on pda or sabouraud agar after 45 days of incubation at 30  c , or blood agar at 37  c .
our patient slowly improved and intravenous ( iv ) methylprednisolone was tapered from 125  mg every 8  h to 40  mg twice a day over a 7 day period . on hospital day 15 , methylprednisolone was discontinued and empiric antifungal treatment was changed from iv amphotericin b to voriconazole 40  mg iv every 12  h. on hospital day 18 , ecmo treatment was terminated , and our patient was weaned successfully from mechanical ventilation and extubated .
she was continued on voriconazole , but developed transaminitis prompting cessation of the drug due to hepatotoxicity .
further antifungal treatment or steroid treatments were not administered . on hospital day 27 , our microbiology laboratory and a state reference laboratory independently identified our mold as aspergillus fumigatus employing phenotypic criteria ( fig .
partial calmodulin , -tubulin , and its gene sequences were obtained via pcr amplification of genomic dna extracted from our clinical isolate .
based on dna sequencing , the mold was identified as the morphologically identical but phylogenetically distinct species neosartorya udagawae ( tables 13 ) . as a result of profound deconditioning , our patient was transferred on day 28 to an inpatient unit to complete intensive physical therapy .
subsequently , she returned to work and had no evidence of recurrent infection on follow - up examination .
neosartorya spp . are the complementary mating types of aspergillus and are required for sex to occur .
although neosartorya infections are uncommon , cases of neosartorya infection misidentified as a. fumigatus are well represented in the literature , suggesting the incidence of neosartorya infections may be higher than suspected . in a retrospective evaluation of specimens banked from patients diagnosed with invasive pulmonary aspergillosis at nih between 2000 and 2008 , vinh et al .  
conducted multilocus dna sequencing to re - examine 36 cases of infection attributed to a. fumigatus based on phenotypic identification . of these , four cases were found to be caused by n. udagawae ; three were seen in patients with chronic granulomatous disease , and one was in a patient with myelodysplastic syndrome .  
the median duration of illness was approximately seven times longer than that typically observed for illness caused by a. fumigatus .
the mics of various antifungals for the n. udagawae isolates were higher than those for a. fumigatus , and the disease caused by n. udagawae was refractory to standard therapy . to make a precise identification of the mold cultured from our patient , dna was extracted and partial its , -tubulin and calmodulin genes were amplified with  touchdown 
pcr with the annealing temperature running from 55 to 45  c , and sequences were generated .
an ncbi blast query of the 798  bp partial -tubulin gene sequence against the ncbi genbank nr / nt nucleotide database returned six n. udagawae isolates with 100% similarity , and returned 28 n. udagawae and seven aspergillus sp . sequences at the 99% similarity level ( table 1 ) .
a query of the 373  bp partial its gene sequence returned seven n. udagawae with 100% similarity ( table 2 ) .
a query of the 743  bp partial calmodulin gene sequence returned five aspergillus sp . with 100% similarity , and 10 n. udagawae and one aspergillus sp . hits at 99% similarity ( table 3 ) . sequences from type strains of species near n. udagawae , high similarity sequences found in the blast search , and the sequences from our clinical isolate were aligned and analyzed with mega 5.2 .
2 ) show large diversity among n. udagawae isolates and reveal that our isolate is in the n. udagawae clade .
the aspergillus sp . sequences with identical or high similarity sequences to n. udagawae represent asexual colonies of this heterothallic species . because the species is heterothallic
the morphology of the aspergillus phase is very similar to a. fumigatus and the definitive identification is through dna sequencing and phylogenetic analysis ( tables 13 ) .
histopathology and culture , ideally in combination , are the current basis for diagnosis and identification of fungal infections .
our patients isolate only produced diagnostic conidiophores after 12 days of incubation , and then it was misidentified as aspergillus fumigatus by two microbiology laboratories .
although it took a week to grow the mold in culture and extract dna to definitively identify the isolate as n. udagawae using its , -tubulin and calmodulin sequence data , the identification could have been done in 23 days if genomic dna had been extracted directly from the original slant culture .
prompt identification of invasive molds directly from clinical specimens is not feasible by conventional microbiologic techniques , but molecular methods for rapid detection of molds directly from bal fluid offer the potential opportunity to direct antifungal treatment and guide evaluation of underlying host susceptibility .
pcr / esi - ms is capable of rapid identification of fungal pathogens directly from clinical specimens .
institutional irb approval was obtained for submission of samples of our patients bal fluid for testing by pcr / esi - ms , as described by shin et al . .
bal fluid from the first bronchoscopy was fixed for cytology testing and unavailable for pcr / esi - ms testing .
pcr / esi - ms was performed on bal fluid from the second bronchoscopy , which was performed on hospital day 7 , 48  h after initiation of amphotericin b treatment .
pcr / esi - ms detected a neosartorya sp . from both rml and left lingular bal fluid specimens .
although airborne conidia of neosartorya sp . are conceivably capable of causing an apba - like syndrome that is seen with a. fumigatus in asthmatic patients , this has not been described .
our patient did not have a history of asthma , nor did her laboratory testing exhibit peripheral eosinophilia or elevated ige titers suggestive of apba .
her response to steroid treatment is consistent with what would be expected in patient with the fibroproliferative stage of ards .
infection has not been previously described . due to uncertainty about the species of mold responsible for the pulmonary infection in our patient
, she was unnecessarily exposed to toxic empiric antifungal treatment with amphotericin b. more timely identification of the pathogen would have resulted in earlier initiation of pathogen specific directed therapy .
while colonization is always a concern with clinical cultures from intubated patients , identification of n. udagawae by gene sequencing of mold grown from multiple lobes in both lungs and by pcr / esi - ms directly from bal fluid samples taken after initiation of antifungal treatment strongly supports the biologic plausibility of n. udagawae as the etiology of ards in our patient .
finally , given that the mics of most antifungals for n. udagawae isolates are higher than for a. fumigatus , and that disease caused by n. udagawae can be refractory to standard therapy , rapid recognition of n. udagawae as the cause of disease can be critical .
presently , it is unclear how often neosartorya udagawae and other species within aspergillus section fumigati cause invasive pulmonary or sinus infections , or allergic bronchopulmonary or sinus manifestations . as molecular modalities such as -tubulin and calmodulin sequencing , and pcr / esi - ms
become integrated into diagnostic clinical microbiology laboratories , detection of putatively less common invasive molds such as neosartorya sp . causing pulmonary disease may increase , thereby providing a more detailed picture of the spectrum of mycoses that infect humans and other animals .
r. sampath and k.s . lowery are salaried employees of ibis biosciences , a division of abbott .
 we describe the first reported case of acute respiratory distress syndrome ( ards ) attributed to neosartorya udagawae infection . 
 this mold grew rapidly in cultures of multiple respiratory specimens from a previously healthy 43-year - old woman . 
 neosartorya spp . 
 are a recently recognized cause of invasive disease in immunocompromised patients that can be mistaken for their sexual teleomorph , aspergillus fumigatus . because the cultures were sterile , phenotypic identification was not possible . 
 dna sequencing of its , calmodulin and -tubulin genes supported identification of neosartorya udagawae . 
 our case is the first report of ards associated with neosartorya sp . infection and defines a new clinical entity . 
Introduction
Case
Discussion
Conflict of interest