Detection and Localization ofRickettsia spin Mealybug

Abstract

Mealybug, Phenacoccus solenopsis Tinsley, is a sap-sucking hemipteran insect. It is an agricultural pest that is now widely distributed in India. In this study we report the presence of Rickettsia from P. solenopsis. We constructed a 16S rRNA gene library to study the bacterial diversity associated with this insect and we found that all the clones from the library were only of Candidatus Tremblaya phenacola. This study also highlights that the normal protocol adopted to study the bacterial diversity from environmental sample, by preparation of a 16S rRNA gene library, does not work when the bacterial population is highly skewed in favor of one bacteria (primary endosymbiont in this case). Hence, we used bacterial genus specific polymerase chain reaction primers to test the presence of any of the widely known secondary endosymbionts associated with insects. We tested for the presence of Cardinium, Rickettsia, Wolbachia, and Arsenophonus in P. solenopsis collected from 10 different locations across India. Only Rickettsia was detected from four locations while we were not able to find any other bacteria. We confirmed the presence of these bacteria by localizing Rickettsia and the primary endosmbiont, Candidatus Tremblaya sp. to the bacteriocyte of P. solenopsis using fluorescent in situ hybridization.

Detection and Localization of Wolbachia in Thrips palmi Karny (Thysanoptera: Thripidae)

“…Genus specific primers have been reported to be more sensitive for the detection of less populated bacterium [21]. We observed an amplification product of 632 bp which was in accordance with the expected size of 16S rRNA gene with genus specific primers of Wolbachia [22] in all three populations. Similarity search result using BLAST algorithm (search was performed against nucleotide sequence nr/nt, scoring parameter; match/mismatch scores were 1/-2 and gap cost set as linear) in NCBI showed that sequence of this product was 99 % identical to previously deposited sequences of Wolbachia from other insects in the GenBank.…”

“…Genomic DNA samples of three populations of well identified T. palmi were subjected to PCR for detection of Wolbachia with genus specific 16S rRNA gene primers. Conventionally, a 16S rRNA gene library is made to study bacterial endosymbiont population, but this technique does not detect bacterial endosymbionts which are rare [22]. Genus specific primers have been reported to be more sensitive for the detection of less populated bacterium [21].…”

“…Such structures have earlier been observed to show auto fluorescence in RNase digested controls and in absence of probe during Rickettsia sp. detection in mealybug [22].…”

An exceptional family: Ophiocordyceps‐allied fungus dominates the microbiome of soft scale insects (Hemiptera: Sternorrhyncha: Coccidae)

“…These discrepancies are possibly due to the use of different probes in each study. The Wolbachia and Rickettsia that were found in high relative abundance in C. floridensis and M. mangiferae , respectively, are likely secondary symbionts of these species, as they often are in other sternorrhynchan families (e.g., Chiel et al., ; Singh, Kumar, Thomas, Ramamurthy, & Rajagopal, ; Zytynska & Weisser, ). Among the many bacterial OTUs that were amplified from our samples, there were also sequences of Uzinura diaspidicola , Buchnera aphidicola , Tremblya princeps and Moranella endobia (Appendix ).…”

Microbial Associates of the Vine Mealybug Planococcus ficus (Hemiptera: Pseudococcidae) under Different Rearing Conditions

“…The evolution of obligate nutritional symbiosis has been highly dynamic in mealybugs, with some groups harboring Tremblaya alone, others harboring Tremblaya and Moranella, and others harboring an as yet unnamed obligate symbiont in the Bacteroidetes [35]. Mealybug bacterial facultative symbionts have received much less attention, although infections with acetic acid-producing bacteria [21], Spiroplasma [24], and Rickettsia [63] have been documented. Little is known about fungal associates of mealybugs although Buchner [6] reported yeast-like symbionts in the mealybug Rastrococcus iceryoides.…”