Increased Constituent Ratios of Klebsiella sp., Acinetobacter sp., and Streptococcus sp. and a Decrease in Microflora Diversity May Be Indicators of Ventilator-Associated Pneumonia: A Prospective Study in the Respiratory Tracts of Neonates
Ventilator-associated pneumonia (VAP) is a common complication and cause of death in neonates on mechanical ventilation. However, it is difficult to define the causes of VAP. To understand the causes of VAP, we undertook a prospective study based on the diversity of the microflora in VAP. The experimental group consisted of newborns who suffered from respiratory distress syndrome (RDS) and VAP, while the control group suffered from RDS without VAP. Sputa were collected within 1, 3, and 5 days of ventilation and were divided into six groups. DNA was extracted from the samples, and the 16S rDNA was PCR amplified, separated using denaturing gradient gel electrophoresis (DGGE), cloned and sequenced. The resulting sequences were compared using BLAST. The DGGE pictures were measured, and the richness, Shannon-Wiener index, and cluster maps were analyzed. No differences were found regarding the constituent ratio of any genus between the Non-VAP and VAP group within 1 day after intubation. After 1 to 3 days, the constituent ratios of Klebsiella sp., Acinetobacter sp., and Streptococcus sp. in the VAP group were higher than those in the Non-VAP group, and the ratios of Serratia sp. and Achromobacter sp. were lower. After 3 to 5 days, the ratios of Klebsiella sp., Acinetobacter sp., Serratia sp., and Achromobacter sp. were lower than those in the Non-VAP group. The richness and Shannon-Wiener index of the Non-VAP group were higher than those of the VAP group from 1 to 3 days after intubation, while no differences were found within 1 day and from 3 to 5 days. We conclude that during the first three days of intubation, the microflora diversity in the lower respiratory tract was reduced due to VAP, and the greater constituent ratios of Klebsiella sp., Acinetobacter sp., and Streptococcus sp. in the sputum may be indicators of VAP.
References
[1]
Michael Balter (2012) Taking stock of the human microbiome and disease. Science 336: 1246–1247. doi: 10.1126/science.336.6086.1246
[2]
Yalaz M, Altun-K？ro？lu O, Ulusoy B, Yildiz B, Akisu M, et al. (2012) Evaluation of device-associated infections in a neonatal intensive care unit. Turk J Pediatr 54: 128–135.
[3]
Grgurich PE, Hudcova J, Lei Y, Sarwar A, Craven DE (2013) Diagnosis of ventilator-associated pneumonia: controversies and working toward a gold standard. Curr Opin Infect Dis 26: 140–150. doi: 10.1097/qco.0b013e32835ebbd0
[4]
Celik IH, Oguz SS, Demirel G, Erdeve O, Dilmen U (2012) Outcome of ventilator- associated pneumonia due to multidrug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa treated with aerosolized colistin in neonates: a retrospective chart review. Eur J Pediatr 171: 311–316. doi: 10.1007/s00431-011-1537-z
[5]
Peters BM, Jabra-Rizk MA, O’May GA, Costerton JW, Shirtliff ME (2012) Polymicrobial Interactions: Impact on Pathogenesis and Human Disease. ASM 25: 193–213. doi: 10.1128/cmr.00013-11
[6]
Kellen berger E (2001) Exploring the unkown. The silent revolution of microbiology. EMBO Rep 2: 5–7.
[7]
Schmid TM, De Long EF, Pace NR (1991) Analysis of marin epicoplankton community by16SrRNA gene clon in gand sequencing. J Bacteriol 173: 4371–4378.
[8]
Handelsman J, Rondon MR, Brandy SF, Clardy J, Goodman RM (1998) Molecular biologic alaccess to the chemistry of unkown soil microbes: anewfrontier for natural products. Chem Biol 5: R245–249. doi: 10.1016/s1074-5521(98)90108-9
[9]
Relman DA, Loutit JS, Schmidt TM (1990) The agent of bacillary angiomatosis. An approach to the identification of uncultured pathogens. N Engl J Med 323: 1573–1580. doi: 10.1056/nejm199012063232301
[10]
Relman DA, Schmidt TM, MacDermott RP, Falkow S (1992) Identification of the uncultured bacillus of Whipple’ s disease. N Engl J Med 327: 293–301. doi: 10.1056/nejm199207303270501
[11]
Cairns S, Thomas JG, Hooper SJ, Wise MP, Frost PJ, et al. (2011) Molecular Analysis of Microbial Communities in Endotracheal Tube Biofilms. PLoS One 6: e14759. doi: 10.1371/journal.pone.0014759
[12]
Craven DE, Hudcova J, Lei Y (2011) Diagnosis of ventilator-associated respiratory infections (VARI): microbiologic clues for tracheobronchitis (VAT) and pneumonia (VAP). Clin Chest Med 32: 547–557. doi: 10.1016/j.ccm.2011.06.001
[13]
Wang F, He B (2013) The role of endotracheal aspirate culture in the diagnosis of ventilator-associated pneumonia: a meta analysis. Zhonghua Jie He He Hu Xi Za Zhi 36: 27–32.
[14]
Payne MS, Goss KC, Connett GJ, Kollamparambil T, Legg JP, et al. (2010) Molecular microbiological characterization of preterm neonates at risk of bronchopulmonary dysplasia. Pediatr Res 67: 412–418. doi: 10.1203/pdr.0b013e3181d026c3
[15]
Wang Y, Hoenig JD, Malin KJ, Qamar S, Petrof EO, et al. (2009) 16S rRNA gene-based analysis of fecal microbiota from preterm infants with and without necrotizing enterocolitis. ISME J 3: 944–954. doi: 10.1038/ismej.2009.37
[16]
Hill TC, Walsh KA, Harris JA, Moffett BF (2003) Using ecological diversity measures with bacterial communities. FEMS Microbiol Ecol 43: 1–11. doi: 10.1111/j.1574-6941.2003.tb01040.x
[17]
Signoretto C, Bianchi F, Burlacchini G, Sivieri F, Spratt D, et al. (2010) Drinking Habits Are Associated with Changes in the Dental Plaque Microbial Community. J Clin Microbiol 48: 347–356. doi: 10.1128/jcm.00932-09
[18]
Cao M, Feng Y, Wang C, Zheng F, Li M, et al. (2011) Functional definition of LuxS, an autoinducer-2 (AI-2) synthase and its role in full virulence of Streptococcus suis serotype 2.. J Microbiol 49: 1000–1011. doi: 10.1007/s12275-011-1523-1
[19]
Ahmed NA, Petersen FC, Scheie A A (2009) AI-2/LuxS is involved in increased biofilm formation by Streptococcus intermedius in the presence of antibiotics. Antimicrob Agents Chemother 53: 4258–4263. doi: 10.1128/aac.00546-09
[20]
Shaffer RK (2013) The Challenge of Antibiotic-Resistant Staphylococcus : Lessons from Hospital Nurseries in the mid-20th Century. Yale J Biol Med 86: 261–270.
[21]
Hamet M, Pavon A, Dalle F, Pechinot A, Prin S, et al. (2012) Candida spp. airway colonization could promote antibiotic-resistant bacteria selection in patients with suspected ventilator-associated pneumonia. Intensive Care Med 38: 1272–1279. doi: 10.1007/s00134-012-2584-2
[22]
Gapon MN, Ternovskaia LN (2011) Biochemical criteria for severity of colonic dysbacteriosis. Klin Lab Diagn 2: 18–20.
[23]
Kuziemski A, Czerniak B, Frankowska K, Gonia E (2012) Molecular epidemiology of Acinetobacter baumannii strains isolated in the years 2008–2010 in University Hospital no.2 in Bydgoszcz. Przegl Epidemiol 66: 403–407.
[24]
Nazik H, Ongen B, Ilktac M, Aydin S, Kuvat N, et al. (2012) Carbapenem resistance due to Bla(OXA-48) among ESBL-producing Escherichia coli and Klebsiella pneumoniae isolates in a univesity hospital, Turkey. Southeast Asian J Trop Med Public Health 43: 1178–1185.
[25]
Bahrani-Mougeot FK, Paster BJ, Coleman S, Barbuto S, Brennan MT (2007) Molecular analysis of oral and respiratory bacterial species associated with ventilator-associated pneumonia. J Clin Microbiol 45: 1588–1593. doi: 10.1128/jcm.01963-06
[26]
Vorwerk C, Manias K, Davies F, Coats TJ (2011) Prediction of severe bacterial infection in children with an emergency department diagnosis of infection. Emerg Med J 28: 948–951. doi: 10.1136/emj.2009.087783
[27]
Chalmers NI, Palmer RJ Jr, Cisar JO, Kolenbrander PE (2008) Characterization of a Streptococcus sp.-Veillonella sp. community micromanipulated from dental plaque. J Bacteriol 190: 8145–8154. doi: 10.1128/jb.00983-08
[28]
Hancock KL, Schermer TR, Holton C, Crockett AJ (2012) Microbiological contamination of spirometers - an exploratory study in general practice. Aust Fam Physician 41: 63–64.
[29]
Pierce-Hendry SA,Dennis J (2010) Bacterial culture and antibiotic susceptibility testing. Compend Contin Educ Vet 32: E1–5; quiz E6.
