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). DNA was prepared from bacterial cultures as described. The DNA was used as a template in PCRs with the primers described on the Bacillus cereus MLST Web site 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. The combination of the 7 alleles determines a given sequence type. A greater number of alleles that match between strains indicates a higher level of relatedness. 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. 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. All other case-patients recovered and were discharged.
Investigation of the ventilation procedures in the NICU
revealed that most equipment used for respiratory care was disposable,
designated for single-patient use. The Draeger Evita v500 ventilator 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. 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. We were able to fully characterize 4 of the 9 isolates. 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 solution and then sent for steam autoclave sterilization at 134°C (273.2°F). 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.
