an immunology graduate student born after the discontinuation of routine smallpox vaccination was working with multiple strains of vaccinia as part of her thesis research . one morning in october 2004 , she noticed the onset of itching , tearing , palpebral swelling , and conjunctival injection in her left eye . viral conjunctivitis was diagnosed by her student health services , and over - the - counter tetrahydrozoline hydrochloride eye drops were prescribed . during the next 4 days , the eye became swollen , red , and painful ; malaise , fatigue , and subjective fever also appeared . on day 5 the patient went to a private ophthalmologist , who referred her to a specialty eye hospital . physical examination at the eye hospital demonstrated a painful left eye with 3 + chemosis in the eyelids and conjunctiva and symblepharon at the lower pole of the eye . a 0.5-cm vesicle was noted above the left canthus ( figure 1 ) . left ocular range of motion , including palpebral motion , was severely limited . computed tomographic scan of the orbits indicated left preseptal cellulitis without evidence of orbital cellulitis . the diagnosis of vaccinia infection was not suspected until examination at the eye hospital , when the student first mentioned her work with vaccinia . contact precautions were then initiated , and a scraping of the vesicle above the canthus was sent to the pennsylvania bureau of laboratories for vaccinia testing . the patient was started on trifluridine and bacitracin ointments , broad - spectrum systemic antimicrobial agents , and pain medication ; she was admitted to the hospital . , additional vesicles appeared on the lower conjunctiva ( figure 2 ) , and periorbital swelling increased . polymerase chain reaction ( pcr ) testing at the pennsylvania bureau of laboratories showed evidence of vaccinia ; results were confirmed at the centers for disease control and prevention ( cdc ) . at this time , vaccinia immune globulin ( vig ) , 6,000 u / kg iv , was administered . less than 24 hours after vig administration , the patient 's pain and swelling were substantially decreased . the patient continued to improve over the next 2 days and was discharged to her home on day 9 . photographer : e. claire newbern . a public health investigation of the patient 's home and work contacts and the research laboratory was initiated . because cutaneous lesions from vaccinia typically appear 35 days after inoculation ( 5 ) , investigators postulated that the patient contracted her infection within the 7 days before symptom onset . the patient was considered infectious from the beginning of this period until hospitalization ( 11 days ) . during this time all contacts were interviewed with a standard questionnaire to ascertain the extent of their contact with the patient , vaccination status , and information about laboratory work practices , if applicable . acute- and convalescent - phase paired serum samples were drawn at 1 and 6 weeks after exposure from all work contacts , as well as from the patient 's closest home contact . paired serum samples from the patient 's consenting contacts were examined for evidence of orthopoxvirus - reactive antibody by using techniques described elsewhere ( 9 ) . immunoglobulin g ( igg ) optical density cutoff value ( cov ) = 0.214408 ;  , not performed . three laboratory workers who also manipulated vaccinia were vaccinated 1 week before the convalescent - phase blood sample was drawn ( workers 2 , 3 , and 4 ) . all 3 had been vaccinated as children ; workers 2 and 3 had orthopoxvirus - reactive igg levels present above the cov . worker 4 did not have igg levels above the cov either before or after her recent vaccination . a laboratory inspection , which included a review of experiments performed by the patient during the week before symptom onset , was conducted . although laboratory staff generally followed established biosafety precautions ( 10 ) , review of laboratory practices showed several opportunities for virus exposure . instances occurred in which samples with low titers of live virus were removed from the biosafety cabinet , transported to other parts of the facility , and manipulated . in addition , laboratory staff routinely vortexed tubes containing live virus outside of the biosafety cabinet . most important , no laboratory workers had been vaccinated in the past 10 years , as recommended by cdc ( 6,10 ) . to identify the specific infecting strain of vaccinia , the virus isolated from the patient 's canthus lesion was sequenced . briefly , a 3.7-kbp amplicon was generated and sequenced from the thymidine kinase region of the viral genome by using the following primers : tkj2r forward 5-acgtg atgga tatat taaag tcgaa and tkj2r reverse 5-gttta tctaa cgaca caaca tcca . amplification was performed with the expand long template pcr kit ( roche molecular biologicals , indianapolis , in , usa ) and a cetus model 9700 thermocycler ( perkin - elmer life and analytical sciences , boston , ma , usa ) at 92c  2 min , followed by 30 cycles of 92c  10 s , 55c  30 s , and 68c  3 min . purified , amplified dna was sequenced with a ceq 8000 genetic analysis system ( beckman - coulter , fullerton , ca , usa ) . sequencing showed that the infecting virus was a unique form of recombinant western reserve vaccinia constructed in the research laboratory and routinely used by the patient ; it had been last used as a control strain during a multiday experiment performed in the 5 days before the patient 's symptoms began . at one point in this experiment , a 96-well plate containing small amounts of live vaccinia  infected mammalian cells was removed from the biosafety cabinet and hand - carried to another room , where the lid of the plate was removed , and the cells were examined for fluorescence . the student did not wear eye protection during this phase of the experiment ; whether she wore gloves is unclear . the investigation of the laboratory and examination of clinical specimens from the patient and contacts enabled investigators to pinpoint the source of infection to a single experiment . during the period when the patient could have become infected , she was the only laboratory member to use the culprit vaccinia strain , and she used it only while performing this particular experiment . lack of seroconversion among the other staff argues against widespread environmental viral contamination in the laboratory . during the time when she could have become infected , the student had also worked with a different strain of vaccinia in titers as high as 110 pfu / ml . however , all of the work with virus at this titer occurred in the biosafety cabinet . although the exact mechanism of infection could not be determined , the location of the principal lesion at the inner canthus suggests either inadvertent inoculation from hand to eye or inoculation through aerosolization of virus ( 5 ) . regardless , both mechanisms indicate that existing biosafety precautions in the laboratory were likely insufficient . biosafety level 2 ( bsl-2 ) precautions are recommended for laboratories and persons who manipulate nonattenuated strains of vaccinia virus ( 10 ) . this recommendation assumes a priori that all such persons will be adequately vaccinated against the virus . however , this report and others of laboratory - acquired vaccinia infections demonstrate that vaccination is being waived in certain institutions ( 13,11 ) . no current recommendations exist in the united states for the level of precautions to be used by unvaccinated personnel . we believe that vaccination would probably have prevented or attenuated this patient 's infection and that it should continue to be recommended for laboratory workers who handle vaccinia . however , given that vaccination has risks of its own that might reduce its use ( including a rate of ocular complications of 1020/1 million immunizations ) ( 5,6,12,13 ) , biosafety recommendations for unvaccinated personnel should be specifically addressed . chiefly intended to protect against agents with potential for respiratory transmission , bsl-3 precautions emphasize protection from exposure to potentially infectious aerosols ( 10 ) . cdc has previously recommended increased biosafety precautions for laboratories with unvaccinated personnel who manipulate monkeypox virus ( 14 ) . implementing certain bsl-3 precautions in this case , e.g. , performing all manipulations of virus in the biosafety cabinet or other enclosed equipment , frequent glove changing accompanied by handwashing , and always wearing goggles or face shields when working with virus outside of a primary containment device , would have minimized the potential for human error and might have prevented this infection . use of eye protection should be particularly stressed , as serious eye infections can occur even in previously vaccinated persons ( 15 ) . no systematic monitoring of vaccinia infection in laboratory workers currently exists , so the full extent of the problem is unknown , vaccination is the best way to prevent or mitigate accidental infection ( 4 ) and should continue to be recommended for laboratory workers handling nonattenuated strains of vaccinia . if vaccination is impossible , workers should implement more stringent biosafety practices , such as consistently using goggles and performing all manipulations of virus in the biosafety cabinet .
we report a case of ocular vaccinia infection in an unvaccinated laboratory worker . 
 the patient was infected by a unique strain used in an experiment performed partly outside a biosafety cabinet . 
 vaccination should continue to be recommended , but laboratories with unvaccinated workers should also implement more stringent biosafety practices .