Ventilator support is essential part of care in neonatal intensive care unit (NICU), but ventilator-associated pneumonia (VAP) can occur as a serious complication. National Nosocomial Infections Surveillance system (NNIS) data (2004) from USA reported pooled mean VAP rate of 1.4-3.5/1000 ventilator days [1]. In developing countries, the reported rates of VAP are significantly higher, ranging from 16.1 to 89 episodes per 1,000 ventilator days [2,3].

Infants who required ventilatory support were intubated by oro-tracheal route in NICU or labour room, and then put on Neonatal Ventilators (Maquet servo-i, Avea standard or Schiller graph NET advance). Disposable ventilator circuits (RT 126 dual limb infant ventilator circuits, Fisher and Paykel) were connected on ventilator through servo-controlled humidifier (Fisher and Paykel MR850). The disposable tubings were not changed till extubation or until visible soiling. Humidifier was filled with sterile water by a closed system. All ventilated babies were nursed in the supine position, and routine care as per NICU protocol was provided. Suctioning and collection of sample for ETA was done by an open suction using Hagedorn method [10], when first suction was required for the presence of secretions. Suctioning was performed by using sterile feeding tube or suction catheter and syringe. If the yield was <1 mL, the procedure was repeated by instilling 0.5 mL of normal saline (drawn from a freshly opened ampoule) into the endotracheal tube. Sample collected in the syringe was sent for quantitative culture and microscopic analysis along with sample of saline taken for suction. Studied infants were carefully followed up for signs of VAP. This included, apart from clinical examination, regular recording of body temperature, ventilator setting (peak inspiratory pressure, positive end expiratory pressure and fractional oxygen (FiO2)), arterial blood gas, leukocyte count, serial C-reactive protein, blood culture and chest radiographs. Blood culture was done as a part of the sepsis work-up profile. No infant received steroid, local or oral antibiotic, H2 blocker or proton pump inhibitor.

A smear was prepared from ETA for Gram staining to determine polymorphonuclear (PMNL) cells and the type of organism. Microscopy was done at high magnification (x400) by using VISION 2000 LED microscope and recorded as: PMNL count: >10/HPF; 5-10/HPF or <5/HPF or Nil /HPF. Smear of ETA was scored as per Bartlett scoring system [11] and bacteria were classified as gram positive or negative; cocci or bacilli. Culture of ETA and ET Tip was done on Blood and Mac Conkey agar. Antibiotic sensitivity was done on Muller Hinton agar and results were expressed as colony forming units/mL. With quantitative analysis of ETA and ET tube tip, the threshold for diagnosing VAP in this study was considered as 10 5CFU/mL [12].

Data analysis was done by calculating mean and standard deviation for continuous data and using unpaired t test/ Mann whitney U test for statistical significance. Pearson’s chi-square test or fisher’s exact test was used for categorical data. P value <0.05 was considered as statistically significant. Statistical analysis was done using the software SPSS version 17.0 for windows

Results

A total of 326 neonates were screened out of which 68 were included in the study (Fig. 1). The baseline demographic data of the study population were comparable (Table I). Twenty-eight (41.2%) neonates developed VAP. Mean (SD) time of ET aspirate sample collection was 53.2 (2.9) h for (SD) microscopy and culture, and of ET tube tip sample was 115.2 (40.8) h. The mean (SD) time of result of endotracheal aspirate microscopy and endotracheal aspirate culture was 55.7 (4.3) h and 108.3 (19.7) h, respectively in comparison to diagnosis made at 143.5 (23.3) h as per CDC criteria (P<0.001).

Fig.1 Flow of patients in the study.

 

TABLE I	Baseline Demographic and Risk Factors in the Study Population

Variable	VAP  (28)	NO VAP  (40)
Age (wks) *	36.9 (1.8)	38 (1.7)
Weight (kg) *	2.3 (0.6)	2.7 (0.5)
Male gender	15 (53.6%)	21 (52.5%)
Meconium stained amniotic fluid	17 (60.7%)	15 (37.5%)
Vaginal delivery	22 (78.6%)	30 (75%)
Resuscitation at birth
Bag and mask ventilation	7 (25.0%)	8 (20%)
Intubation	13 (46%)	6 (15%)
APGAR at 5 minutes <7	19 (67.9%)	11 (27.5%)
No. of endotracheal tube changes*	1.82 (0.67)	1.65 (0.66)
Duration of ventilation*	14.07 (2.23)	10.18 (2.02)
Mortality (all cause)	9 (32.1%)	10 (25%)
*values in mean (SD); VAP: Ventilator-associated pneumonia.

The sensitivity, specificity, negative predictive value and positive predictive value of ETA microscopy (³5 PMNL/HPF and >10 PMNL/HPF), ETA culture (colony count >105 CFU/mL), endotracheal tube tip culture (colony count >105 CFU/mL) are shown in Table II. In our study, organisms cultured in ETA were Acinetobactor (29.4%), Klebsella (4.4%), Pseudomonas (3%), MRSA (3%); mixed growth was seen in one case.

TABLE II  Results of Microscopy and Culture

In this study, ETA microscopy (³ 5PMNL/HPF) and ETA culture colony count >105 CFU/mL were found to be useful for early diagnosis of VAP with significantly lower diagnosis time. No significant complications of ETA were found in this study.

The ETA culture sensitivity and specificity when cut off was >105 CFU/mL for diagnosis of VAP in our study was comparable to findings noted by Labenne, et al. [6] where samples were retrieved by BAL technique. The mean time to diagnosis of VAP as per CDC criteria in our study was comparable to that documented by Tripathi, et al. [3], but detection of VAP could be done earlier by us using ETA microscopy and culture.

We conclude that ETA culture colony count (>105 CFU/mL) and ETA microscopy ³5PMNL/HPF is supportive in the objective diagnosis of VAP with added advantage of early diagnosis.

1. National Nosocomial Infections Surveillance System Report: Data summary. Am J Infect Control. 2004;32:470-85.

2. Foglia E, Meier M, El-ward A. Ventilator-associated pneumonia in neonatal and pediatric intensive care units. Clin Microbiol Rev. 2007;20:409-25.

3. Tripathi S, Malik GK, Jain A. Study of ventilator associated pneumonia in neonatal intensive care unit: characteristics, risk factors and outcome. Internet J Med Update. 2010;5:12-9.

4. Garner JS, Jarvis WR, Emori TG, Horan TC, Hughes JM. CDC definitions for nosocomial infections, 1988. Am J Infect Control. 1988;16:128-40.

5. Luna CM, Vujacich P, Niederman M. Impact of BAL data on the therapy and outcome of ventilator-associated pneumonia. Chest. 1997;111:676-85.

6. Labenne M, Poyart C, Rambaud C, Goldfarb B, Pron B, Jouvet P,　et al. Blind protected specimen brush and bronchoalveolar lavage in ventilated children. Crit Care Med.　1999;27:2537-43.

7. Gauvin F, Dassa C, Chaibou M, Proulx F, Farrell CA, Lacroix J. Ventilator-associated pneumonia in intubated children: comparison of different diagnostic methods.　Pediatr Crit Care Med.　2003;4:437-43.

8. Hagedorn MI, Gardner SL, Abman SH: Respiratory diseases; Handbook of Neonatal Intensive Care. St. Louis: Mosby 1989. 381.

9. Bartlett JG, Ryan KJ, Smith TF. Laboratory Diagnosis of Lower Respiratory Tract Infection. In: Washington JA ed, Cumitech 7A, Washington DC: American Society for Microbiology.1987. p. 1-18.

10. Marquette CH, Georges H, Wallet F, Ramon P, Saulnier F, Neviere R, et al. Diagnostic efficiency of endotracheal aspirates with quantitative bacteria cultures in intubated patients with suspected pneumonia. Comparison with the protected specimen brush. Am Rev Respir Dis. 1993;148:138-44.