Intraspecific and interspecific comparison among bacterial communities in salivary glands and gut of five Culicoides spp. (Diptera: Ceratopogonidae), vectors of bluetongue virus

Bacteria within the vector species can impact various physiological traits, including likely digestion, nutrition, survivability, and fecundity. Bacterial communities within insects vary across tissues and alter depending on diet regime, host taxa, and parity. Salivary glands and the gut are two significant organs for pathogen replication and dissemination. Despite various records on viruses, protozoa, and nematodes, data on bacterial communities within salivary glands (SG) and gut (GT) are lacking within the five vectors of bluetongue virus (BTV), Culicoides actoni Smith, C. fulvus Sen & Das Gupta, C. imicola Kieffer, C. jacobsoni Macfie and C. orientalis Macfie. It is well documented that certain bacteria can impact on vectorial capacity of viruses of biting midges. The culturable bacterial composition is investigated in SG and GT across sexes and age-graded females of five field-collected species to unveil interspecific and intraspecific variation among the bacteria associated with vector species across sexes and different organs. Bacterial colonies were identified by biochemical tests and Sanger sequencing of the partial 16S rRNA gene. A total of 114 bacterial isolates, of which 61 and 53 isolates were retrieved from SG and GT of five species, respectively. The isolates belonged to the phyla Firmicutes and Proteobacteria. Bacillus spp. was the most abundant, followed by Lysinibacillus spp. The relative abundances of bacterial genera identified from five species across sexes and organs are as follows: SG- i) C. actoni: Bacillus (11.47 %), Lysinibacillus (1.63 %), Paenibacillus (4.91 %), Enterococcus (1.63 %), Serratia (1.63 %); ii) C. fulvus: Bacillus (11.47 %), Enterococcus (3.27 %); iii) C. imicola: Bacillus (13.11 %), Brevibacillus (1.63 %), Paenibacillus (3.27 %), Enterococcus (3.27 %); iv) C. jacobsoni: Bacillus (11.47 %), Paenibacillus (1.63 %), Enterococcus (4.91 %) and v) C. orientalis: Bacillus (13.11 %), Paenibacillus (3.77 %), Pseudomonas (1.88 %) and GT- i) C. actoni: Bacillus (7.54 %), Lysinibacillus (9.43 %), Paenibacillus (1.88 %), Serratia (1.88 %); ii) C. fulvus: Bacillus (9.43 %), Lysinibacillus (1.88 %), Cytobacillus (1.88 %), Pseudomonas (1.88 %); iii) C. imicola: Bacillus (16.98 %), Lysinibacillus (5.66 %), Brevibacillus (1.88 %); iv) C. jacobsoni: Bacillus (15.09 %) and v) C. orientalis: Bacillus (18.86 %), Brevibacillus (3.77 %), Pseudomonas (1.88 %). The bacteria exhibiting beta haemolytic activities were: 5 Bacillus spp., 3 Lysinibacillus spp., Cytobacillus kochii, Brevibacillus parabrevis, E. faecium, Paenibacillus lautus, S. marcescens, Pseudomonas stutzeri, while 5 Bacillus spp., and L. mangiferihumi were gamma haemolytic. Bacteria of the SG were identified for the first time, along with the haemolytic microbiota of these species. This repository of haemolytic gut bacteria may be targeted further to detect their probable roles in vivo blood meal digestion. Our findings record the occurrence of the following pathogenic bacteria: Bacillus cereus, Brevibacillus parabrevis, L. fusiformis, S. marcescens, and P. stutzeri within SG, which may be transmitted to their hosts through blood feeding.