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\begin{document}

\title{Culture Result Discrepancy Between Laboratories for Nontuberculous
Mycobacteria in Patients with Cystic Fibrosis}



\author[1]{Mariana Bedoya}%
\author[2]{Max Salfinger}%
\author[3]{Alejandra Weisman}%
\author[3]{Andrew Colin}%
\affil[1]{Monroe Carell Junior Children's Hospital at Vanderbilt}%
\affil[2]{University of South Florida Morsani College of Medicine}%
\affil[3]{University of Miami School of Medicine}%


\vspace{-1em}



  
  \date{June 17, 2024}


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\selectlanguage{english}
\begin{abstract}
NTM-infected CF patients may be at risk of being underdiagnosed or
inappropriately treated when relying on culture and susceptibility
results from non-specialized laboratories. Since this was a small study
with convenience samples, a larger study needs to be carried out. If our
findings are confirmed, the drivers should be elucidated for the
discrepant results. Given the increasing prevalence of NTM in the
population at large and not in the CF community
alone{[}{[}13{]}{]}(\#ref-0013), elucidating any differences in testing
to ensure the correct identification, including subspeciation and
antimicrobial susceptibilities should be paramount%
\end{abstract}%



\sloppy


To the Editor:

\textbf{Culture Result Discrepancy Between Laboratories for
Nontuberculous Mycobacteria in Patients with Cystic Fibrosis}

\textbf{Mariana Bedoya, MD1, Max Salfinger, MD2, Alejandra Weisman 3,
Andrew Colin, MD3}

1 Division of Allergy, Immunology, Pulmonary and Sleep Medicine, Monroe
Carell Jr. Children's Hospital at Vanderbilt, Nashville, TN 2.
University of South Florida, College of Public Health \& Morsani College
of Medicine, Tampa, FL 3. Division of Pediatric Pulmonology, Miller
School of Medicine, University of Miami, Miami, FL

Nontuberculous mycobacteria (NTM) are ubiquitous organisms that can
cause severe chronic pulmonary infection, particularly in patients with
cystic fibrosis (CF), and have become a rising concern worldwide.{[}1{]}
The predominant NTM species found among people with CF
include\emph{Mycobacterium avium} and \emph{Mycobacterium abscessus}
.{[}2{]} The prevalence of NTM based on large studies in adult and
pediatric patients with CF in the Americas, Europe, and Australia varies
between 6-13\%.{[}3-6{]} In a large study in the United States (US), NTM
prevalence was shown to vary significantly among patients with CF by
geographic area, an observation attributed to environmental
factors.{[}7{]} Another US study identified significant spatial
clustering of NTM in 8 states, including Florida.{[}8{]}

The 2018 CF patient registry reported an annual prevalence of 13.6\% of
positive cultures of one or more mycobacterial species isolated from
those who had a mycobacterial culture {[}9{]}, and the longitudinal
prevalence (years 2010-2016) of one or more NTM species isolated was
19\%.{[}10{]} We led a Florida-based, two-center prevalence review in
conjunction with University of Florida in 2012, and found 8 out of 85
patients (9.4\%) under 18 years of age had at least one positive culture
of NTM among the two CF centers.{[}11{]} Our pediatric practice in
Miami, FL, has been experiencing a dramatic increase in the number of CF
patients with at least one culture positive for NTM; our prevalence in
2017 was 31.8\%.

NTM is essentially undiagnosable from conventional throat
cultures.{[}11{]} NTM require adequate samples of sputum or
bronchoalveolar lavage (BAL) fluid. Specialty laboratories are key for
reliable culture results. It is crucial to identify the sample as
deriving from a CF patient for the laboratory, especially from those who
harbor \emph{Pseudomonas} species, since sample decontamination is
essential to prevent overgrowth of bacteria that are omnipresent in CF.
These bacteria could interfere with the growth of NTM, resulting in
false negative results.{[}12{]} Subsequent to growth, identification to
species and subspecies level and antimicrobial susceptibility testing,
including drug resistance markers, is also crucial. These tests will
determine whether patients will be placed on prolonged, complex, and
onerous antimicrobial regimens. Currently, at our institution, acid-fast
bacilli (AFB) sputum cultures are sent to commercial laboratories when
obtained in our outpatient practice. The commercial laboratory is
dictated by insurance, without flexibility in the choice of specialty
laboratories. Inpatient samples (sputum and BAL) are analyzed in our
teaching hospital laboratory, where a positive AFB culture will trigger
PCR for \emph{Mycobacterium tuberculosis.} If negative
for\emph{Mycobacterium tuberculosis} , the sample is forwarded to a
national reference laboratory for culture, speciation and antimicrobial
susceptibility testing.

The ominous reality of the tripling prevalence of NTM in our pediatric
CF center was particularly concerning for several patients who presented
with deteriorating health, deemed disproportionate to their known
infectious etiologies and in whom we excluded other specific potential
comorbidities such as allergic bronchopulmonary aspergillosis,
malnutrition, and CF related diabetes. While following guidelines for
annual AFB cultures, we suspected false negative AFB cultures or
possibly reporting an incorrect identification or antimicrobial
susceptibility results, all of which can affect diagnosis and outcome.

Our goal was to ascertain the reliability of diagnostic tests in our
teaching hospital laboratory (A) and referral commercial laboratories
(B) and compare their reported results to a specialized laboratory for
NTM (C).

We obtained 19 samples (BAL and/or sputum) for AFB from patients with
suspected or known NTM. Identical samples were contemporaneously sent to
laboratory A and/or B, and all were sent to laboratory C (Table 1). All
cultures were followed for up to 56 days. One sample was lost in
shipping to laboratory C and removed from analysis. Eight of 18 samples
grew NTM in at least one laboratory. Fifty percent of the positive
samples for NTM had discrepancies. Two samples (11.1\% of total) sent to
laboratory A or laboratory B failed to grow NTM when compared to
laboratory C, and 2 samples (11.1\% of total) had growth at both
laboratory B and laboratory C but revealed different species.
Additionally, laboratory B failed to report antimicrobial susceptibility
testing results in three out of four of its positive
samples\textbf{.}Overall, we found discrepancies in 22.2\% of AFB
culture results from CF patients from duplicate BAL or sputum samples
between laboratory A and/or laboratory B compared to the specialized
laboratory for NTM (C).

We conclude that NTM-infected CF patients may be at risk of being
underdiagnosed or inappropriately treated when relying on culture and
susceptibility results from non-specialized laboratories. Since this was
a small study with convenience samples, a larger study needs to be
carried out. If our findings are confirmed, the drivers should be
elucidated for the discrepant results. Given the increasing prevalence
of NTM in the population at large and not in the CF community
alone{[}13{]}, elucidating any differences in testing to ensure the
correct identification, including subspeciation and antimicrobial
susceptibilities should be paramount.

Table 1\selectlanguage{english}
\begin{longtable}[]{@{}lllllll@{}}
\toprule
& \textbf{Patient Initials} & \textbf{Date} & \textbf{Sample} &
\textbf{A} & \textbf{B} & \textbf{C}\tabularnewline
\midrule
\endhead
\textbf{1} & \textbf{JD} & 2/2/2017 & Sputum & NT & \emph{M.
avium-intracellulare} complex & \emph{M. abscessus} subsp.
\emph{massiliense} \& \emph{M. chimaera}\tabularnewline
\textbf{2} & \textbf{LP} & 2/2/2017 & Sputum & NT & No growth & \emph{M.
abscessus} subsp. \emph{massiliense}\tabularnewline
\textbf{3} & \textbf{AZ} & 2/22/2017 & Sputum & No growth & NT & No
growth\tabularnewline
\textbf{4} & \textbf{DL} & 5/4/2017 & BAL & No growth & NT & No
growth\tabularnewline
\textbf{5} & \textbf{DP} & 11/4/2017 & Sputum & NT & \emph{M.
chelonae-abscessus} & \emph{M. abscessus} subsp.
\emph{massiliense}\tabularnewline
\textbf{6} & \textbf{DC} & 4/17/2017 & BAL & No growth & NT & No
growth\tabularnewline
\textbf{7} & \textbf{BB} & 5/15/2017 & BAL & No growth & NT & No
growth\tabularnewline
\textbf{8} & \textbf{LP} & 5/24/2017 & Sputum & ~NT & No growth & No
growth\tabularnewline
\textbf{9} & \textbf{NG} & 12/15/2017 & Sputum & \emph{M. abscessus} &
~NT & \emph{M. abscessus} subsp. \emph{massiliense}\tabularnewline
\textbf{10} & \textbf{DL} & 7/24/2018 & BAL & No growth & ~NT & No
growth\tabularnewline
\textbf{11} & \textbf{DP} & 7/16/2018 & Sputum & NT & \emph{M.
avium-intracellulare~}complex & \emph{M. abscessus} subsp.
\emph{abscessus}\tabularnewline
\textbf{12} & \textbf{NG} & 2/8/2018 & Sputum & No growth & NT & No
growth\tabularnewline
\textbf{13} & \textbf{JD} & 9/8/2018 & BAL & No growth & NT & No
growth\tabularnewline
\textbf{14} & \textbf{MR} & 8/16/2018 & BAL & No growth & No growth & No
growth\tabularnewline
\textbf{15} & \textbf{MK} & 8/16/2020 & Sputum & ~NT & \emph{M.
abscessus} & \emph{M. abscessus} subsp.
\emph{massiliense}\tabularnewline
\textbf{16} & \textbf{NG} & 1/16/2019 & BAL & No growth & No growth &
Lost sample\tabularnewline
\textbf{17} & \textbf{DL} & 8/2/2019 & Sputum & No growth & NT &
\emph{M. abscessus} subsp. \emph{massiliense}\tabularnewline
\textbf{18} & \textbf{DC} & 4/3/2020 & BAL & No growth & NT & No
growth\tabularnewline
\textbf{19} & \textbf{MK} & 6/23/2020 & BAL & \emph{M. abscessus} & NT &
\emph{M. abscessus} subsp. \emph{massiliense}\tabularnewline
\bottomrule
\end{longtable}

A: Teaching hospital laboratory; B: Referral commercial laboratories; C:
Specialized laboratory for nontuberculous mycobacteria; BAL:
Bronchoalveolar lavage; M.: \emph{Mycobacterium;} subsp.: subspecies;
NT: Not tested

\textbf{References}

1. Skolnik, K., G. Kirkpatrick, and B.S. Quon, \emph{Nontuberculous
Mycobacteria in Cystic Fibrosis.} Curr Treat Options Infect Dis,
2016.\textbf{8} (4): p. 259-274.

2. Floto, R.A., et al., \emph{US Cystic Fibrosis Foundation and European
Cystic Fibrosis Society consensus recommendations for the management of
non-tuberculous mycobacteria in individuals with cystic
fibrosis.}Thorax, 2016. \textbf{71 Suppl 1} : p. i1-22.

3. Adjemian, J., K.N. Olivier, and D.R. Prevots, \emph{Epidemiology of
Pulmonary Nontuberculous Mycobacterial Sputum Positivity in Patients
with Cystic Fibrosis in the United States, 2010-2014.} Ann Am Thorac
Soc, 2018. \textbf{15} (7): p. 817-826.

4. Aiello, T.B., et al., \emph{Prevalence and clinical outcomes of
nontuberculous mycobacteria in a Brazilian cystic fibrosis reference
center.} Pathog Dis, 2018. \textbf{76} (5).

5. Gardner, A.I., et al., \emph{Epidemiology of Nontuberculous
Mycobacteria Infection in Children and Young People With Cystic
Fibrosis: Analysis of UK Cystic Fibrosis Registry.} Clin Infect Dis,
2019. \textbf{68} (5): p. 731-737.

6. Roux, A.L., et al., \emph{Multicenter study of prevalence of
nontuberculous mycobacteria in patients with cystic fibrosis in
france.}J Clin Microbiol, 2009. \textbf{47} (12): p. 4124-8.

7. Adjemian, J., K.N. Olivier, and D.R. Prevots, \emph{Nontuberculous
mycobacteria among patients with cystic fibrosis in the United States:
screening practices and environmental risk.} Am J Respir Crit Care Med,
2014. \textbf{190} (5): p. 581-6.

8. Adjemian, J., et al., \emph{Spatial clusters of nontuberculous
mycobacterial lung disease in the United States.} Am J Respir Crit Care
Med, 2012. \textbf{186} (6): p. 553-8.

9. \emph{Cystic Fibrosis Foundation Patient Registry Annual Data
Report.} 2018.

10. Cystic-Fibrosis-Foundation-Patient-Registry. \emph{Cystic Fibrosis
Foundation Patient Registry Annual Data Report} . 2017.

11. Yousef S, S.P., Baker D, Della Volpe A, Colin A, Quizon
A,\emph{Prevalence of non-tuberculous mycobacteria (NTM) in pediatric
patients with cystic fibrosis (CF) in Florida -- A two center
study.}Paediatr Respir Rev, 2013. \textbf{14} (Supp 2): p. S72.

12. Griffith, D.E., et al., \emph{An official ATS/IDSA statement:
diagnosis, treatment, and prevention of nontuberculous mycobacterial
diseases.} Am J Respir Crit Care Med, 2007. \textbf{175} (4): p.
367-416.

13. Ratnatunga, C.N., et al., \emph{The Rise of Non-Tuberculosis
Mycobacterial Lung Disease.} Frontiers in Immunology, 2020. \textbf{11}
.

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