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at an sbv information meeting , 60 shepherds > 18 years of age were recruited for this study . after obtaining written informed consent , we administered a standardized questionnaire .
we collected information about age , sex , sbv infection in their livestock , exposure to sick lambs , frequency of insect bites , personal health , and categories of signs of disease after exposure .
we developed an indirect fluorescent antibody test ( ifat ) for primary testing of human serum .
conjugated secondary antibodies against sbv - specific igm or igg ( antibovine for positive control ) were used . for the ifat
, all heat - inactivated serum specimens were tested in dilutions of 1:20 and 1:80 on glass slides with noninfected and sbv - infected vero cells .
an sbv antibody  positive serum sample from an experimentally infected cow was used as a positive control . to check for background signals and possible cross - reactivity ,
a serum neutralization test ( snt ) was developed for confirmation of indeterminate and positive results .
serial dilutions of the test serum ( lowest dilution 1:5 ) were incubated for 1 h at 37c with an equal volume of cell culture supernatant containing 100 infectious doses of sbv and subsequently mixed with vero cells . to detect sbv - specific rna
, we performed a 1-step real - time reverse transcription quantitative pcr ( rt - qpcr ) on serum , as described ( 1 ) .
all 60 participants ( 75% male ; median age 48 years [ interquartile range ( iqr ) 4156 years ] ) reported sheep husbandry in the sbv - epizootic area ( table 1 ) .
altogether , 48 ( 80% ) participants had contact with lambs that had characteristic malformations or with the respective birth products ( median 10 [ iqr 420 ] sick lambs ) . in livestock from 36 ( 60% ) participants , sbv was laboratory confirmed .
median time from first signs in animals to blood withdrawal was 45 days ( iqr 3966 days ) .
a total of 55 ( 98% ) of 56 participants self - reported insect bites during late summer to autumn ; among these , 22 ( 39% ) indicated frequent insect bites .
nine ( 15% ) shepherds reported having had signs and symptoms since the disease had appeared in the study area or after handling diseased animals ( table 1 ) : myalgia and arthralgia ( 7 shepherds ) , headache ( 4 ) , fever ( 4 ) , skin rash ( 2 ) , and respiratory problems ( 2 ) .
of the 36 shepherds whose livestock had laboratory - confirmed sbv infection , 5 ( 14% ) reported signs and symptoms : myalgia and arthralgia ( 4 shepherds ) , headache ( 2 ) , fever ( 2 ) , skin rash ( 2 ) , and respiratory problems ( 2 ) .
self - reported signs and symptoms of fever , headache , skin rash , myalgia / arthralgia , respiratory problems , or photophobia since sbv infection appeared in the study area or after handling diseased animals and resulting from unknown cause in each exposure category . 
no sbv - specific antibodies were detected in any serum specimens ( table 2 ) .
eight specimens showed indeterminate fluorescent signals in the ifat at a 1:20 serum dilution for igg ( n = 1 ) or igm ( n = 7 ) but were not reactive at 1:80 ( figure ) .
these 8 samples were retested by snt ( serum dilution 1:10 ) and showed no virus inhibition at any serum dilution during 7 days of incubation .
two ( 25% ) of these 8 shepherds reported symptoms . for the bovine control serum ,
* sbv , schmallenberg virus ; ifat , indirect fluorescent antibody test ; snt , serum neutralization test ; rt - qpcr , quantitative reverse transcription pcr . 
performed in only 8 serum samples with ifat indeterminate results ; 2/8 ( 25% ) reported symptoms as outlined in table 1 .
fluorescent light microscopy images of serum samples tested for antibodies to schmallenberg virus by indirect fluorescent antibody test on infected vero cells mixed with noninfected vero cells .
a ) nonreactive negative serum ; b ) positive serum reactive with infected cells only ( green ) ; c ) indeterminate serum with faint nonspecific reactivity .
we investigated the risk for human infection after possible high exposure to an emerging vector - borne epizootic disease through contact with infected animals and tissues or through insect bites .
no evidence of sbv infection among the shepherds was found by molecular and serologic tests , even though most of the shepherds had received substantial exposure through repeated direct contact with sheep with laboratory - confirmed sbv - infection and with birth products known to contain high virus loads in the sbv - epizootic area .
reported symptoms were compatible with illnesses commonly experienced during the winter ( i.e. , influenza - like illness caused by human respiratory viruses ) without considerable differences between the exposure categories . the likelihood of virus detection by rt - qpcr is certainly limited because sbv viremia in livestock lasts only a few days ( 4,8 ) .
however , after the end of the viremic phase , detection of specific antibodies can be expected .
the period between exposure and sampling was sufficiently long to for antibodies to have developed after infection .
furthermore , a large proportion of the participants indicated having been frequently bitten by insects in the epizootic area .
midge bites are difficult to recall , and therefore this exposure could not be assessed precisely .
although sbv has been isolated from certain midge species , entomologic knowledge about the ability of different midge species to transmit sbv , i.e. , vector competence and host feeding behavior , is still scarce .
nevertheless , on the basis of results from our study and the phylogenetic relationship of sbv , we conclude that the novel virus is unlikely to pose a threat to humans by transmission from infected livestock or from midges .
schmallenberg virus , a novel orthobunyavirus , is spreading among ruminants , especially sheep , throughout europe . 
 to determine the risk for human infection , we conducted a survey among shepherds to assess possible exposure and symptoms . 
 we also performed serologic and molecular assays . 
 no evidence of transmission to humans was detected .