tissue specimens were sent to the national reference center ( marseilles , france ) either frozen or in transport media . csd was definitively diagnosed when a specimen was positive for 2 genes of bartonella spp ( 1 ) . total genomic dna was extracted from samples by using a qiaamp tissue kit ( qiagen , hilden , germany ) . before 2005 , pcr amplification and sequencing of the internal transcribed spacer ( its ) region and pap31 gene were used for detecting b. henselae and thus confirming csd ( 1 ) . beginning in 2005 , real - time pcr to amplify the its region and pap31 gene was used ( 8) . for all assays , dna from b. elizabethae and b. henselae houston - i was used as the positive control for the its region and the pap31 gene , respectively ( 1 ) . to exclude false - positive results , we performed a second independent extraction when false - positive or unexpected results were obtained . results were confirmed by using pcr amplification and sequencing aimed at 16s rrna gene ( 8) . epi info version 6.0 software ( centers for disease control and prevention , atlanta , ga , usa ) was used for significance variations in the number of positive specimens between 2 consecutive months , nonconsecutive months , and seasons ( p<0.05 ) . the mantel - haenszel test or the fisher exact test positive csd cases were plotted for each month ( figure 1 ) to identify seasonal distributions of csd from 1999 through 2009 . monthly mean numbers of csd were lowest from may through august , followed by significant increases during august september ( p = 0.002 ) and during november  december ( p = 0.01 ) . during december  january , the mean number of csd cases decreased significantly ( p = 0.005 ) , then plateaued from january through march ( table ) . cases decreased slightly in april , then decreased significantly during april  may ( p = 0.002 ) . mean numbers of patients tested for cat - scratch disease and for whom the disease was diagnosed , france , 19992009 . the odds of a csd diagnosis based on lymph node biopsy was 9.2 higher in december , the month with the highest number of cases ( 92/493 ) , than in july , the month with the lowest number of cases ( 12/493 ) . the number of csd cases was significantly higher in autumn ( october  december ) than in summer ( july  september ) ( p<0.0001 ) . fewer cases were identified in winter ( january  march ) than autumn ( p = 0.02 ) , and cases decreased significantly from winter to spring ( april  june ) ( p = 0.0001 ) . the number of cases did not differ significantly from spring to summer ( p = 0.06 ) . our findings that the number csd cases in france varies by season are similar to findings in japan and the united states . in japan , 64% of csd occurred during september  december and peaked in november ( 7 ) . in the united states , most csd have occurred during the last 6 months of the year , with a peak in september ( 9 ) . moreover , the analysis of 3 us national databases indicated that most csd cases have occurred during september  january , with peaks in november and december ( 5 ) . on the other hand , 60% of admissions for csd in children in the united states have occurred during july  october ( 6 ) . the fact that the united states is a large country with diverse climates , whereas continental france has a more homogeneous climate , may explain the differences in seasonality . the presence of ct . felis fleas is essential for maintaining b. henselae infection within the cat population ( 2 ) . flea infestation is more frequent in bacteremic than in nonbacteremic cats , particularly in pet cats ( 10 ) . after adult cat fleas parasitize a host cat , they feed on its blood and transmit b. henselae . fleas go through 4 life cycle stages : egg , larva , pupa , and imago ( adult ) ( figure 2 ) . temperature and relative humidity are the 2 most essential factors for the successful reproduction , development , and survival of fleas ( 11 ) . cats reported to have been infested with fleas during the preceding 6 months were more likely than cats without fleas to be seropositive ( 12 ) , and the seroprevalence of b. henselae is higher in the pet cat population in warm , humid climates than in cold , dry climates because ct . felis fleas are more common in warmer climates ( 10 ) . as a result , cats have more fleas during the summer and autumn months than in the other 2 seasons ( 13 ) . explanation of cat scratch disease seasonality by seasonality of the birth of cats and the activity of their fleas , france , 19992009 . 53% of 94 stray cats were infected with either b. henselae or b. clarridgeiae ( 14 ) . in paris , chomel et al . reported a b. henselae seroprevalence of 36% in 64 pet cats , of which 11% were b. henselae infected ( 12 ) . determined that 16.5% of cats tested were bartonella infected , and 41% were seropositive for b. henselae or b. clarridgeiae ( 15 ) . risk for bartonella infection or seropositivity was higher in cats from multicat households and in cats adopted from the pound or from the street ( 15 ) . feline sexual activity also may influence the seasonality of csd . in the northern hemisphere , cat reproduction increases during spring and summer , and kittens stay with their mothers until they are 1216 weeks of age . b. henselae infection appears to be more common in young cats ( 10 ) , and infection decreases with the length of cat ownership ( 15 ) . in addition , cats encounter more fleas during summer and autumn , and transmission of b. henselae from cat to cat is facilitated during this period ( 13 ) . in conclusion , csd is a seasonal disease in france with increased incidence in autumn , with peaks in december , and a decrease in spring . this pattern may be explained by seasonality in cat reproductive behavior , their fleas activities , and the fact that during summer cats spend most time outside the house , whereas during autumn they stay indoors .