Huanglongbing (HLB) (yellow shoot disease) is a highly destructive disease that threatens citrus production worldwide. The disease was first observed in Guangdong, P.R. China, over 100 years ago, and was found in Florida, United States, in 2005. 'Candidatus Liberibacter asiaticus' has been associated with HLB in many citrus-growing regions around the world, including Guangdong and Florida. The global epidemiology of HLB, as well as management of the disease, relies on knowledge of 'Ca. L. asiaticus' populations in different geographical regions around the world. In this study, we identified a genetic marker containing small tandem repeats in the genome of 'Ca. L. asiaticus' and comparatively analyzed the tandem repeat numbers (TRNs) in 'Ca. L. asiaticus' populations from Guangdong and Florida. Analyses of TRNs showed that the bacterial population in Guangdong was different from that in Florida. The Guangdong population consisted predominately of strains with a TRN of 7 (TRN(7)) at a frequency of 47.6%. The Florida population consisted predominately of strains with a TRN of 5 (TRN(5)) at a frequency of 84.4%. TRNs ranged from 3 to 16. The apparent absence of TRNs of 9, 10, 11, and 12 separated the bacterial strains into two groups: TRNs < 10 (TRN(<10)) and TRNs > 10 (TRN(>10)). In Florida, TRN(<10) strains (103/109, or 94.5%) were widely distributed in all HLB-affected counties. TRN(>10) strains (6/109, or 5.5%) were found in central Florida. This is the first report documenting the differentiation of 'Ca. L. asiaticus' populations between Asia and North America and the possible presence of two differentially distributed genotypes of 'Ca. L. asiaticus' in Florida.

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Polymerase chain reaction (PCR) primers were designed and tested for identification of immature parasitoids in small grain cereal aphids and for estimation of parasitism rates. PCR technique was evaluated for 1) greenhouse-reared greenbugs, Schizaphis graminum (Rondani), parasitized by Lysiphlebus testaceipes Cresson and 2) aphids collected from winter wheat fields in Caddo County, Oklahoma. For greenhouse samples, parasitism frequencies for greenbugs examined by PCR at 0, 24, and 48 h after removal of L. testaceipes parasitoids were compared with parasitism frequencies as determined by greenbug dissection. PCR was unable to detect parasitism in greenbugs at 0 and 24 h postparasitism, but it was able to detect parasitoids 48 h after parasitoid removal at frequencies that were not significantly different from dissected samples. Field-collected samples were analyzed by rearing 25 aphids from each sample and by comparing parasitoid frequencies of mummies developed and PCR performed on another 50 aphids. Aphid samples included corn leaf aphids, Rhopalosiphum maidis (Fitch); bird cherry-oat aphids, Rhopalosiphum padi (L.); English grain aphids, Sitobion avenae (F.); and greenbugs. Mummies were isolated until adult emergence, whereupon each parasitoid was identified to species (L. testaceipes was the only parasitoid species found). Parasitism detection frequencies for PCR also were not statistically different from parasitism frequencies of reared aphids. These results indicate that PCR is a useful tool for providing accurate estimates of parasitism rates and especially for identification of immature parasitoids to species.

"Candidatus Liberibacter asiaticus" (CaLas) is associated with citrus Huanglongbing (HLB, yellow shoot disease), which is highly destructive to world citrus production. Understanding the relationships of CaLas isolates from different geographical regions is important for HLB research and development of disease management strategies. In this study, 301 CaLas isolates [85 Brazil, 132 China, and 84 U.S. (83 Florida and 1 California)] were collected, and genomic variations among them were evaluated based on the analyses of two genomic loci: trn1, characteristic of variable tandem repeat numbers (TRNs), and snp1, characteristic of single nucleotide polymorphisms (SNPs). Locus trn1 revealed the homogeneity of all Brazilian isolates, and locus snp1 revealed the homogeneity of most Florida isolates. When the two loci were analyzed simultaneously, i.e., double-locus (DL) analyses, CaLas isolates were clustered mostly according to geographical origins. DL genotype 1 included 97 % of the Chinese isolates, DL genotype 2 included all Brazilian isolates, and DL genotype 3 included 93 % of the U.S. isolates. DL analyses successfully revealed inter-continental overlapping or movement pattern of CaLas isolates. The isolate recently found in California belonged to Asiatic DL genotype 1.