the missouri department of health and senior services conducted this investigation in response to the hospital s identification of an increased number of tracheal aspirates that were positive for b. cereus collected from newborns who were on ventilators during march  may , 2011 . all tracheal aspirate culture results obtained in the neonatal intensive care unit ( nicu ) during january 2010june 2011 were reviewed . nicu data was also searched for positive b. cereus culture from other specimens , such as blood , body fluids , or tissues . investigators thoroughly evaluated respiratory management practices in the unit by direct observation , respiratory records review , and an interview with the respiratory therapist . several environmental cultures were obtained from the flow sensors of the unit s ventilators over the 1-month period . b. cereus isolates were forwarded to the centers for disease control and prevention to be molecularly characterized by using multilocus sequence typing ( mlst ) ( 10 ) . the dna was used as a template in pcrs with the primers described on the bacillus cereus mlst web site ( www.pubmlst.org/bcereus ) for the 7 loci which define the mlst scheme . the sequences for the loci glpf , gmk , ilvd , pta , pur , pyca , and tpi were then assigned allele designations . a greater number of alleles that match between strains indicates a higher level of relatedness ( 10 ) . prevalence of b. cereus  positive specimens was compared by using the mann - whitney u test . retrospective analysis of tracheal aspirate culture results showed significant increase ( p = 0.039 ) in b. cereus isolation between march and may , 2011 ( figure 1 ) . no bacillus spp . were isolated from blood , other body fluids , or tissues during the study period . the chart review of the case - patients comprising the cluster of b. cereus colonization revealed that none received a diagnosis of clinical b. cereus infection . all patients were treated with vancomycin or tobramycin , or both , for indications not related to b. cereus in tracheal aspirate . one case - patient died 108 days later without evidence that b. cereus contributed to the outcome . epidemiologic curve of bacillus spp.positive tracheal aspirates from newborns on ventilators , january 2010january 2012 . investigation of the ventilation procedures in the nicu revealed that most equipment used for respiratory care was disposable , designated for single - patient use . telford , pa , usa ; www.draeger.us/sites/enus_us/pages/hospital/evita-xl.aspx ) was used for mechanical ventilation of infants who were intubated to treat severe respiratory compromise . the draeger evita v500 is a microprocessor controlled ventilator offering both mandatory and spontaneous ventilation modes for adult , pediatric , and neonatal patients . heated and humidified gas flows from the ventilator unit , through the inspiratory circuit and neoflow air flow sensor to the patient through an endotracheal tube . upon exhalation , gas flows back through the air flow sensor into the expiratory circuit and returns to the ventilator through the expiratory flow sensor and exhalation valve . in addition to the ventilator , reusable respiratory equipment comprised a proximal air flow sensor , expiratory flow sensor , exhalation valve , and circuit temperature probe . the sensor closest to the newborn s mouth was an air flow sensor located inside the disposable ventilation circuit ( figure 2 ) . from 9 environmental cultures obtained from 9 air flow sensors , 1 was positive for bacillus spp . , and was later confirmed as b. cereus by the state public health laboratory . mlst was performed for 8 b. cereus isolates from case - patients and for 1 environmental isolate from the air flow sensor . one locus for the remaining 5 strains did not yield an amplicon for sequencing after repeated attempts and , thus , could not be assigned a sequence type . the isolates that included sequence type ( st ) 73 and st94 were closely related to each other because they differed by merely 1 locus , gmk . the strains that were not fully typed because of the inability to obtain sequences for locus pta were also closely related to st73 or st94 because the other loci matched . there was 1 match between strains isolated from 1 case - patient and the air flow sensor , which was st73 . the contaminated air flow sensor was then sterilized by using a steam autoclave . a repeat culture of this sensor after sterilization was negative . we found that air flow sensors were routinely disinfected by placing them in a container with 70% alcohol solution for 60 minutes . after discovery of the air flow sensor contaminated with b. cereus , the disinfection policy was changed . all air flow sensors were first soaked in enzol enzymatic detergent ( asp , irvine , ca , usa ; www.aspjj.com/us/products/enzol ) solution and then sent for steam autoclave sterilization at 134c ( 273.2f ) . after implementation of new disinfection and sterilization procedures , no new cases of b. cereus tracheal colonization were identified in the nursery . in this cluster , contaminated proximal air flow sensors were the likely source of tracheal colonization with b. cereus in newborn infants , supported by a genetic match by mlst between a strain isolated from 1 case - patient and the contaminated air flow sensor . b. cereus transmission from contaminated respiratory equipment has been reported in other geographic areas . in the netherlands , an outbreak of b. cereus infections in a pediatric intensive care unit caused by contaminated reusable ventilator air flow sensors switching to disposable air flow sensors stopped colonization with b. cereus in that unit . in canada , an outbreak of b. cereus infections among patients in an adult icu was linked to colonized ventilator circuitry ( 8) . in the united kingdom , reusable ventilator circuits were also identified as the cause of a b. cereus outbreak among intubated nicu patients ( 9 ) . our investigation underscores the necessity of close monitoring of occurrences of bacillus spp . in tracheal aspirates since clustering of such cases , b. cereus isolates were either st73 , st94 , or closely related to those sequence types . st73 and st94 are associated with strains previously described as having caused illness in elderly persons . strains with st73 were implicated in cases of septicemia ( 12 ) , and of sepsis and pneumonia ( 13 ) . strains with st 94 were recovered from patients with pneumonia ( 14 ) . b. cereus strains harboring b. anthracis plasmids such as pxo1 , have also been associated with severe and fatal respiratory infections ( 15 ) . all case - patients in our investigation were considered to be colonized with b. cereus without clinical implications . since all of them received intravenous antimicrobial drugs effective against b. cereus , it is conceivable that the clinical course of those patients could have been different without such treatment . should not be routinely viewed as clinically insignificant and further testing to determine exact strain should be considered under appropriate clinical and epidemiologic circumstances . proper disinfection of the entire ventilator circuit as recommended by the equipment manufacturer is crucial in avoiding potentially lethal b. cereus infections .
we investigated bacillus cereus  positive tracheal aspirates from infants on ventilators in a neonatal intensive care unit . 
 multilocus sequence typing determined a genetic match between strains isolated from samples from a case - patient and from the air flow sensor in the ventilator . changing the sterilization method for sensors to steam autoclaving stopped transmission .