Endosymbiont-derived metabolites are essential for tick host reproductive fitness.

Ticks, like other obligatory blood-feeding arthropods, rely on endosymbiotic bacteria to supplement their diet with B vitamins lacking in blood. It has been suggested that additional metabolites such as L-proline may be involved in this nutritional symbiosis, but this has yet to be tested. Here, we studied the metabolite-based interaction between the brown dog tick Rhipicephalus sanguineus (Acari: Ixodidae) and its Coxiella-like endosymbionts (CLE). We measured amino acid titers and tested the effect of B vitamins and L-proline supplementation on the fitness of CLE-suppressed female ticks, displaying low titers of CLE. We found higher titers of L-proline in the symbiont-hosting organs of unfed ticks and in engorged blood-fed whole ticks. Supplementation of B vitamins increased the hatching rate of CLE-suppressed ticks; this effect appears to be stronger when L-proline is added. Our results indicate that L-proline is produced by CLE, and we suggest that CLE is essential in states of high metabolic demand that affects tick reproductive fitness, such as oogenesis and embryonic development. These findings demonstrate the broader effect of nutritional symbionts on their hosts and may potentially contribute to the control of ticks and tick-borne diseases. IMPORTANCE: Coxiella-like endosymbionts (CLE) are essential to the brown dog tick Rhipicephalus sanguineus for feeding and reproduction. This symbiosis is based on the supplementation of B vitamins lacking in the blood diet. The involvement of additional metabolites has been suggested, but no experimental evidence is available as yet to confirm a metabolic interaction. Here, we show that B vitamins and L-proline, both of which contribute to tick reproductive fitness, are produced by CLE. These findings demonstrate the importance of symbiont-derived metabolites for the host's persistence and shed light on the complex bacteria-host metabolic interaction, which can be channeled to manipulate and control tick populations.

Metabolomic changes associated with acquired resistance to Ixodes scapularis.

Guinea pigs repeatedly exposed to Ixodes scapularis develop acquired resistance to the ticks (ATR). The molecular mechanisms of ATR have not been fully elucidated, and partially involves immune responses to proteins in tick saliva. In this study, we examined the metabolome of sera of guinea pigs during the development of ATR. Induction of components of the tyrosine metabolic pathway, including hydroxyphenyllactic acid (HPLA), were associated with ATR. We therefore administered HPLA to mice, an animal that does not develop ATR, and exposed the animals to I. scapularis. We also administered nitisinone, a known inhibitor of tyrosine degradation, to another group of mice. The mortality of I. scapularis that fed on mice given HPLA or nitisinone was 26 % and 72 % respectively, compared with 2 % mortality among ticks that fed on control animals. These data indicate that tick bites alter the guinea pig metabolome, and that the tyrosine metabolism pathway can potentially be targeted for I. scapularis control.

Laboratory Management of Mammalian Hosts for Ixodes scapularis -Host-Pathogen Interaction Studies.

Due to their hematophagous life cycle, hard-bodied ticks including the genus Ixodes are a potential vector for numerous pathogenic organisms including bacteria, protozoa, viruses, and infectious prions. The natural geographic range of several hard tick species, includig Ixodes scapularis, has expanded over recent decades. Consequently, there is an ongoing need to maintain, feed, and propagate ticks for host-pathogen interaction studies to better understand and mitigate their impact on human and animal health. Artificial membrane feeding of hard ticks has advanced in recent years, has study design advantages, and should be used, when possible, to reduce animal use, but it also has several limitations that require the continued use of mammalian hosts including mice, guinea pigs, and rabbits. In this overview, we discuss the best management practices for these relevant species with respect to biosafety, health, and optimal host comfort when used in studies that depend on tick feeding. The capsule-jacket method is preferred over the ear sock-E-collar method of tick feeding on rabbit hosts because of better host health, comfort, and increased study versatility.

Salp14 epitope-based mRNA vaccination induces early recognition of a tick bite.

Repeated exposure of animals to Ixodes scapularis ticks can result in acquired tick resistance (ATR). The first manifestation of ATR is erythema at the tick bite site, however, the specific peptide targets and mechanisms associated with this early aspect of ATR are not understood. In this study, we immunized guinea pigs with a lipid nanoparticle containing the mRNA encoding 25 amino acids in the carboxyl terminus of Salp14 (Salp14-C mRNA-LNP), an I. scapularis salivary protein. The animals produced high titers of IgG directed at the carboxyl terminus of Salp14. Guinea pigs immunized with Salp14-C mRNA-LNP and then exposed to I. scapularis, developed erythema at the tick bite site. Transcriptomics of the skin of guinea pigs at the I. scapularis bite sites elucidated selected pathways, including histamine activation, that are associated with the development of erythema. The study demonstrates that an mRNA vaccine encoding a small peptide can induce the initial phase of ATR in guinea pigs.

Metabolomic changes associated with acquired resistance to Ixodes scapularis.

Guinea pigs repeatedly exposed to Ixodes scapularis develop acquired resistance to the ticks (ATR). The molecular mechanisms of ATR have not been fully elucidated, and partially involve immune responses to proteins in tick saliva. In this study, we examined the metabolome of sera of guinea pigs during the development of ATR. Induction of components of the tyrosine metabolic pathway, including hydroxyphenyllactic acid (HPLA), were associated with ATR. We therefore administered HPLA to mice, an animal that does not develop ATR, and exposed the animals to I. scapularis . We also administered nitisinone, a known inhibitor of tyrosine degradation, to another group of mice. The mortality of I. scapularis that fed on mice given HPLA or nitisinone was 26% and 72% respectively, compared with 2% mortality among ticks that fed on control animals. These data indicate that metabolic changes that occur after tick bites contribute to ATR.

Comparative Proteomics of Coxiella like Endosymbionts (CLEs) in the Symbiotic Organs of Rhipicephalus sanguineus Ticks.

Maternally transmitted obligatory endosymbionts are found in the female gonads as well as in somatic tissue and are expected to provide missing metabolite to their hosts. These deficiencies are presumably complemented through specific symbiotic microorganisms such as Coxiella-like endosymbionts (CLEs) of Rhipicephalus ticks. CLEs are localized in specialized host tissue cells within the Malpighian tubules (Mt) and the ovaries (Ov) from which they are maternally transmitted to developing oocytes. These two organs differ in function and cell types, but the role of CLEs in these tissues is unknown. To probe possible functions of CLEs, comparative proteomics was performed between Mt and Ov of R. sanguineus ticks. Altogether, a total of 580 and 614 CLE proteins were identified in Mt and Ov, respectively. Of these, 276 CLE proteins were more abundant in Mt, of which 12 were significantly differentially abundant. In Ov, 290 CLE proteins were more abundant, of which 16 were significantly differentially abundant. Gene Ontology analysis revealed that most of the proteins enriched in Mt are related to cellular metabolic functions and stress responses, whereas in Ov, the majority were related to cell proliferation suggesting CLEs function differentially and interdependently with host requirements specific to each organ. The results suggest Mt CLEs provide essential nutrients to its host and Ov CLEs promote proliferation and vertical transmission to tick progeny. IMPORTANCE Here we compare the Coxiella-like endosymbionts (CLEs) proteomes from Malpighian tubule (Mt) and the ovaries (Ov) of the brown dog tick Rhipicephalus sanguineus. Our results support the hypothesis that CLEs function interdependently with host requirements in each of the organs. The different functional specificity of CLE in the same host suggest that metabolic capabilities evolved according to the constrains imposed by the specific organ function and requirements. Our findings provide specific CLE protein targets that can be useful for future studies of CLE biology with a focus on tick population control.

Biosynthetic Potential of Streptomyces Rationalizes Genome-Based Bioprospecting.

Streptomyces are the most prolific source of structurally diverse microbial natural products. Advancing genome-based analysis reveals the previously unseen potential of Streptomyces to produce numerous novel secondary metabolites, which allows us to take natural product discovery to the next phase. However, at present there is a huge disproportion between the rate of genome reports and discovery of new compounds. From this perspective of harnessing the enduring importance of Streptomyces, we discuss the recent genome-directed advancements inspired by hidden biosynthetic wealth that provide hope for future antibiotics.

Syntrophic association of termite gut bacterial symbionts with bifunctional characteristics of cellulose degrading and polyhydroxyalkanoate producing bacteria.

The guild between higher termites and their partnership with the diverse community of bacteria and archaea in their gut is a marvel evolutionary achievement. Sustained attempts were made worldwide with a quest for identifying viable important biological macromolecule polyhydroxyalkanoate (PHA) accumulating bacteria. Termite gut serve as a novel source for bacteria with dual properties like PHA production as well as cellulose degradation. Among 40 isolates cultivated, 32.5% turned positive for PCR based screening of PhaC gene. The 16S rRNA gene sequencing revealed that elite PHA producer and cellulose degrader which is phylogenetically affiliated to Bacillus cereus. The PHA production was maximized by employing different carbon and nitrogen sources along with altered pH and temperatures. GC-MS, FTIR and 1HNMR analyses confirmed the presence of PHA and the thermal characterization was performed through TGA and DSC for the termite gut isolate. Our results indicated that the combined integrative approach using isolated strains from termite gut would be preferable choice in producing biomolecules from cellulosic materials.