case 1 involved a male neonate delivered by cesarean section in obstetric hospital a in taiwan in april 2013 , after an uneventful pregnancy of 38 weeks . he was fed infant formula in the hospital s nursery . on postpartum day 7 , he had a fever of 39c and tachypnea and was taken to a tertiary hospital .
chest radiograph showed a ground - glass opacity in the middle lobe of his right lung , which subsequently developed into cavities .
the case was reported to the unknown pathogen discovery / investigation group at the taiwan centers for disease control for extensive etiologic survey .
multiplex real - time reverse transcription pcr showed sputum and blood specimens to be negative for adenovirus , respiratory syncytial virus , coronaviruses ( 229e , oc43 , nl63 , hku1 , and middle east respiratory syndrome coronavirus ) , metapneumovirus , influenza virus , parainfluenza viruses 14 , herpes simplex viruses 1 and 2 , varicella - zoster virus , epstein - barr virus , cytomegalovirus , human herpesviruses 6 and 7 , bocavirus , parvovirus , enterovirus , and rhinovirus .
water specimens from all sources with which the patient had contact were tested for legionella by culture .
l. pneumophila serogroup 4 was isolated from 3 tap water sources , and l. pneumophila serogroup 5 was isolated from the cold water source of the hot and cold water dispenser from which water for making infant formula was collected ( table ) .
the water dispenser was located in the room next to the nursery . the hot water faucet provided boiled water at 95100c , and the cold water faucet provided unboiled water that was treated by a built - in reverse osmosis device .
the pulsed - field gel electrophoresis ( pfge ) profiles of the l. pneumophila serogroup 5 strains isolated from a sputum sample from the infant and those from a water sample from the water dispenser were indistinguishable ( figure ) . both strains were sequence type 1032 ( 12 ) . * both case - patients were delivered by cesarean section and were not breast fed or placed in incubators .
st , sequence type ; tmtc , too many to count ; nd , testing not done . 
the first samples from the environments of case - patients 1 and 2 were collected 33 d and 23 d after birth , respectively .
the second samples were collected 34 d and 26 d after birth , respectively . 
concentration of legionella spp . detected in water specimens , attributed to the presence of sampling swabs . 
the pulsed - field gel electrophoresis profile of the strain was indistinguishable from that of the strain isolated from the patient s sputum .
the st of the strain was the same as that of the strain isolated from the patient s sputum .
pulsed - field gel electrophoresis patterns for legionella pneumophila isolates from neonates with legionella infection and hospital water sources .
genomic dna was digested with sfii and separated in 1% agarose gel by bio - rad chef mapper .
lane m , reference size maker ( xbai - digested genomic dna fragments of salmonella enterica ser .
braenderup h9812 ) ; lanes 1 and 2 , clinical and environmental isolates of l. pneumophila serogroup 5 from case - patient 1 ; lanes 3 and 4 , clinical and environmental isolates of l. pneumophila serogroup 1 for case - patient 2 .
parents of the 175 neonates that were delivered 3 months before the patient s birth in hospital a were interviewed by telephone ; 3 of the 175 infants had fever within the first month of life .
serologic testing 3 weeks after their fever episodes was negative for igm or igg antibodies against l. pneumophila serogroups 16 .
the water dispenser in hospital a was replaced with one that provided only hot water .
the point - of - use filters were mounted on the water faucets in the nursery ( pall - aquasafe water filter , portsmouth , uk ) .
no legionella infection was identified in any neonate who was born after the patient s birth in hospital a during the following 8 months .
case 2 involved an asymptomatic male neonate delivered by cesarean section in obstetric hospital b in november 2013 after an uneventful pregnancy of 38 weeks .
he remained asymptomatic and was fed infant formula throughout his 8-day stay in the hospital . at home on the day of discharge
he had tachypnea and cyanosis of the lips during the following days , and on day 10 after birth , he was taken to the same tertiary hospital as case - patient 1 .
bilateral pulmonary infiltrates and mild right pneumothorax were identified . the patient s urine and sputum specimens tested positive for l. pneumophila serogroup 1 .
testing of related environmental water specimens for legionella spp . by culture was negative for all specimens except that from the cold water source of the hot and cold water dispenser in hospital b , which was positive for l. pneumophila serogroup 1 ( table ) .
the water dispenser in the nursery , which consisted of a tank containing boiled , hot water and another tank containing cool water pipelined from the boiled water tank , was used for making infant formula .
the pfge profiles of the strains from the patient and those from the water dispenser were indistinguishable ( figure ) . both strains were sequence type 1 .
the parents of the 79 neonates born 3 months before the patient s birth in hospital b were interviewed by telephone .
the water dispenser in hospital b was replaced with one that provided only hot water .
no legionella infection was identified in any neonate who was born after the patient s birth in hospital b during the following 3 months .
we report 2 cases of neonatal legionellosis associated with infant formula prepared with contaminated water . only the cold water from both hospitals hot and cold water dispensers used for making infant formula was positive for l. pneumophila , with indistinguishable pfge profiles and the same sequence types as those isolated from the neonates .
some environmental specimens were tested twice to compensate for inadequate sensitivity of the culture method and yielded the same results .
no humidifier was used in either case . in the hospital where the first case - patient was identified ,
the water dispenser was not located in the nursery , which likely reduced the risk of transmitting legionella spp . through contaminated aerosol droplets generated by the water dispenser .
aspiration is a major transmission mode in hospital - acquired legionella infection among adults ( 13,14 ) .
neonatal legionella infection through aspiration of contaminated water during water birth has been reported ( 9,10 ) .
our results also indicate the legionella infection described here was likely transmitted by aspiration of contaminated infant formula during the first days of life .
both water dispensers were colonized with legionella spp . although the pervasiveness of these 2 problematic types of water dispenser is not known , hot and cold water dispensers are used in many hospitals in taiwan .
we speculate the complicated pipeline system and inappropriate maintenance of water dispensers might increase the risk for legionella colonization , thus facilitating legionella infection .
most cases were hospital acquired , and the patients had severe pneumonia that was associated with a high mortality rate ( 8,15 ) .
the manifestation of legionella infection is not pathognomonic ( 15 ) , therefore , underdiagnoses or delayed diagnoses combined with inappropriate treatment likely contribute substantially to mortality rates and severity of neonatal legionellosis .
the 2 neonates described here survived after prompt diagnosis and treatment enabled by an extensive etiologic investigation and high suspicion of legionella infection .
our report underscores the importance of suspecting and testing for legionella infection when neonates have respiratory symptoms .
 we report 2 cases of neonatal legionella infection associated with aspiration of contaminated water used in hospitals to make infant formula . 
 the molecular profiles of legionella strains isolated from samples from the infants and from water dispensers were indistinguishable . 
 our report highlights the need to consider nosocomial legionellosis among neonates who have respiratory symptoms . 
The Cases
Conclusion