Sample Selection- Vignettes

Transcription

1 Sample Selection- Vignettes Rangaraj Selvarangan, BVSc, PhD, D(ABMM) Professor, UMKC School of Medicine Director, Microbiology, Virology and Molecular Infectious Diseases Laboratory Director, Laboratory Medicine Research Department of Pathology and Laboratory Medicine Children s Mercy Kansas City

2 Sample Selection- Vignettes Case Presentation: Neonatal HSV disease- HSV PCR for plasma and mucosal surface swabs Adenovirus testing in Immunocompromised patients

3 Neonatal HSV Case History: A 21-day-old baby girl born via uncomplicated vaginal delivery presented to our hospital with several days of vesicular lesions developing around his umbilicus. His mother also noted concurrent maternal genital and oral vesicular lesions concerning for HSV infection. Patient was admitted overnight and started on acyclovir. Plan: Infectious disease was consulted -Acyclovir 20 mg/kg IV q.8 hours started for 14 days of IV therapy -CSF with low white blood cell count normal protein count, CSF HSV PCR negative -HSV PCR obtained from ocular, skin lesion, and anus. The lesion was HSV PCR positive -HSV PCR on plasma was requested.

4 Neonatal HSV disease Neonatal HSV Disease: Skin-Eye-Mucous membrane disease (SEM): 45% CNS disease: 30%- CNS infections and mucocutaneous lesions Disseminated disease: 25% Visceral organ systems involved (liver, lung and most often CNS) Laboratory Testing: Mucosal surface swabs from mouth, nasopharynx, conjunctivae and anus ( surface cultures ) for HSV culture and if desired for HSV PCR assay Specimens of skin vesicles for HSV culture and if desired for PCR CSF sample for HSV PCR assay Whole blood sample for HSV PCR assay Whole blood sample for measuring alanine aminotransferase (ALT). 30 th Edition: Red Book 2015 report of the Committee on Infectious Diseases

5 Poll: HSV plasma PCR

6 Poll: HSV PCR turnaround time

7 Study Design: Retrospective chart review in Seattle Children's Hospital between HSV PCR results from plasma (n=47), CSF (n=56) or both (n=40) at the time of diagnosis were available from 63 infants; 26 with skin/eye/mouth(sem),18 with central nervous system and 19 with disseminated disease (DIS). RESULTS: Plasma HSV PCR was positive in 78% SEM, 64% CNS and 100% DIS disease. Mean plasma viral level was 2.8 log10 copies/ml in SEM, 2.2 log10 copies/ml in CNS, and 7.2 log10 copies/ml in DIS infants. The HSV levels were higher among infants who died compared with surviving infants, 8.1 log10 copies/ml (range ) vs 3.8 log10 copies/ml (range ), P =.001, Level of HSV DNA in the cerebrospinal fluid or in plasma did not correlate with neurologic outcome. Dynamics of HSV clearance from plasma during high-dose acyclovir treatment showed single-phase exponential decay with a median viral half-life of 1.26 days (range: ).

8 HSV PCR and Surface Culture Results for neonatal HSV

9 Melvin et al J Pediatrics 2015; 166 (4):

10 HSV plasma viral clearance in six infants with DIS disease during acyclovir treatment Melvin et al J Pediatrics 2015; 166 (4):

11 HSV plasma PCR HSV plasma PCR was the most frequently positive test with 83% infants testing positive (CSF PCR, plasma PCR and culture). Plasma PCR was the only positive test in 5/47 (11%): 2/18 SEM, 1/11 (CNS) and 2/18 (DIS). In disseminated disease infants with serial viral levels, ACV had a uniform and highly predictable effect on plasma HSV regardless of baseline viral level at initiation of therapy Authors suggest that in the context of clinical worsening, increase in plasma levels may be indication to restart, change antiviral drug or dose. HSV plasma levels at diagnosis are highly correlated with clinical presentation and mortality 2, but not neurological outcome 1. Melvin et al J Pediatrics 2015; 166 (4): Kimura et l J Med Virol 2002; 67:

12 Conclusions HSV PCR testing of the blood specimens can be a useful adjunct to diagnostic evaluation of neonatal HSV infection, especially in infants who present without cutaneous lesions. HSV blood PCR result should not be used to determine extent/classification of disease and thus duration of treatment. Example: Children clinically classified SEM disease and a positive HSV blood PCR would not warrant reclassification as disseminated HSV disease. James and Kimberlin. Journal of Pediatrics 2015: (166)

13 Poll: HSV mucosal surface swabs

14 Adenovirus in Immunocompromised Patients Case Presentation: 6 year old girl with history of hepatoblastoma status post liver transplant one year ago and currently on immunosuppression. He was admitted for a 3 day history of diarrhea, lethargy and hypokalemia. He has had multiple respiratory infections and was most recently admitted for RSV bronchiolitis. He had been doing well at home until yesterday when he developed diarrhea with 2-3 stools in the morning. He is lethargic and has had decreased PO intake. Bacterial stool cultures, C. difficile, and O&P testing were ordered. Physician calls the lab for adenovirus testing of stool sample and requests your input

15 Poll: Adenovirus Stool testing for IC patients

16 Implementation of adenovirus PCR assay Viral surveillance in the immunocompromised (IC) patients: Our laboratory offers quantitative PCR assay for EBV (whole blood), CMV (plasma), BK (plasma, urine) and HHV6 (plasma) viruses on a batched mode. 2 times each week. Implement a Qualitative viral panel surveillance assay for IC patients: Improve TAT, validate broad range of specimen types, reduce send out charges Developed qualitative adenovirus PCR- 5 days/week, plasma, stool, respiratory samples Future Plan: Comprehensive qualitative viral panel surveillance assay for IC patients with improved TAT

17 Poll: Qualitative viral surveillance panel

18 Adenovirus stool detection Adenovirus was detected in our patient stool sample. Adenovirus detection in stool specimen of IC patients is a risk factor for adenoviremia, so adenovirus testing of blood specimen is recommended In our patient, blood viral load for adenovirus revealed 40K copies/ml which climbed to 800K copies/ml a week later. Patient was started on cidofovir therapy and cleared infection two weeks later.

19 Adenovirus stool PCR in IC patients STUDY DESIGN: Retrospective review of adenovirus stool PCR viral loads to determine cutoff for prediction of adenoviremia, in children who underwent an allogeneic hematopoietic stem cell transplant. RESULTS: 117 patients, of which 71 (60%) had diarrhea. Adenovirus was detected in the stool in 39 of 71 (55%) patients. Viral load >10 6 copies/g stool predicted adenoviremia with a sensitivity and specificity of 82%. Median time to viremia detection following stool detection above cutoff was 6.5 days (0-25 days); median blood viral load was 2.6 (<2-7) log 10 copies/ml All patients with adenoviremia had adenovirus detection in stool and adenoviremia did not precede stool detection Srinivasan et al PIDJ 2015