Analysis of the genetic diversity of Phalaenopsis orchids with single nucleotide polymorphisms and snap markers derived from the Pto gene

The Pto gene is a plant gene that has been reported to be involved in resistance to bacterial pathogens. A partial genomic sequence corresponding to Pto (~449 bp) was isolated from 16 species and four hybrids of Phalaenopsis during 2017 at the Department of Agronomy and Horticulture, IPB University, Bogor, Indonesia. Multiple sequence analysis was performed to find putative single nucleotide polymorphisms (SNPs) and design the corresponding single nucleotide-amplified polymorphism (SNAP) markers, which were in turn used to estimate the genetic diversity of 25 Phalaenopsis species. In total, 20 SNPs, of which 14 were nonsynonymous, were identified from the partial Pto sequences. Eighteen SNAP primers were then developed based on these 14 nonsynonymous and four synonymous SNPs. Validation results showed that 15 SNAP primers showed a polymorphism information content exceeding 0.3, suggesting the existence of more than two alleles for this locus. Upon their use, the SNAP markers described 86% of all interspecies variability. The Pto 52, Pto 349, Pto 229, and Pto 380 SNAP markers were very informative in the determination of genetic diversity. Notably, the existence of these nonsynonymous SNPs implied the possibility of functional changes within the amino acid sequence of the putative PTO protein. Thus, the resulting differences in the activity of the PTO protein may be used to breed tolerance to pathogen infection. Further work may be required to establish a functional link between tolerance to pathogens and the presence of Pto-SNAP markers in Phalaenopsis properly.

Molecular Analysis and Phytochemical Profiling to Explore the Diversity of Garcinia Species Collected from Western Ghats of India

Hydroxycitric acid made the genus Garcinia economically important. Genetic and chemical diversity has been studied in Garcinia species using molecular markers, HCA and antioxidant activity. Nine species were collected and screened for molecular diversity and six were subjected to analyse antioxidant and HCA content and its interspecies variability. A total of 129, 125 and 89 bands with polymorphism of 78.74%, 78.4% and 93.36% were obtained using ISSR, RAPD and EST-SSR, respectively. The average PIC value obtained with ISSR, RAPD and EST-SSR markers was 0.9161, 0.9440 and 0.8903, respectively. Determined HCA content by HILIC-HPLC system using 0.1% orthophosphoric acid and acetonitrile (30:70) as mobile phase in fruit powder of various Garcinia species was found to be significantly different. G. gummi-gutta, G. indica and G. xanthochymus are rich of HCA containing 12.44±1.04%, 7.92±0.83% and 6.3±0.286%, respectively. G. morella, G. talbotii and G. celebica contained very negligible amount of HCA, 0.023±0.012%, 0.083±0.034% and 0.34±0.013%, correspondingly. G. talbotii showed high antioxidant capacity (95.40±0.720). Below that G. indica and G. xanthochymus were showing significant amount of total phenols (1.23±0.015 and 1.07±0.008), flavonoids (11.17±0.075 and 12.35±0.219) and antioxidant activity (90.73±0.976 and 91.37±0.854). Correlation analysis found significant association between molecular and chemical variation indicating influence of genetic background on the observed HCA and antioxidant profiles. The conducted analysis showed the most distinct species at the genetic and chemical levels were G. gummi-gutta, G. indica and G. xanthochymus. This study signifies the utility of molecular and chemical fingerprints for commercial exploitation of HCA from Garcinia species.

High prevalence and diversity of Bartonella in small mammals from the biodiverse Western Ghats

Bartonella species are recognized globally as emerging zoonotic pathogens. Small mammals such as rodents and shrews are implicated as major natural reservoirs for these microbial agents. Nevertheless, in several tropical countries, like India, the diversity of Bartonella in small mammals remain unexplored and limited information exists on the natural transmission cycles (reservoirs and vectors) of these bacteria. Using a multi-locus sequencing approach, we investigated the prevalence, haplotype diversity, and phylogenetic affinities of Bartonella in small mammals and their associated mites in a mixed-use landscape in the biodiverse Western Ghats in southern India. We sampled 141 individual small mammals belonging to eight species. Bartonella was detected in five of the eight species, including three previously unknown hosts. We observed high interspecies variability of Bartonella prevalence in the host community. However, the overall prevalence (52.5%) and haplotype diversity (0.9) was high for the individuals tested. Of the seven lineages of Bartonella identified in our samples, five lineages were phylogenetically related to putative zoonotic species–B. tribocorum, B. queenslandensis, and B. elizabethae. Haplotypes identified from mites were identical to those identified from their host species. This indicates that these Bartonella species may be zoonotic, but further work is necessary to confirm whether these are pathogenic and pose a threat to humans. Taken together, these results emphasize the presence of hitherto unexplored diversity of Bartonella in wild and synanthropic small mammals in mixed-use landscapes. The study also highlights the necessity to assess the risk of spillover to humans and other incidental hosts.

Wnt5A-Mediated Actin Organization Regulates Host Response to Bacterial Pathogens and Non-Pathogens

Wnt5A signaling facilitates the killing of several bacterial pathogens, but not the non-pathogen E. coli DH5α. The basis of such pathogen vs. non-pathogen distinction is unclear. Accordingly, we analyzed the influence of Wnt5A signaling on pathogenic E. coli K1 in relation to non-pathogenic E. coli K12-MG1655 and E. coli DH5α eliminating interspecies variability from our study. Whereas cell internalized E. coli K1 disrupted cytoskeletal actin organization and multiplied during Wnt5A depletion, rWnt5A mediated activation revived cytoskeletal actin assembly facilitating K1 eradication. Cell internalized E. coli K12-MG1655 and E. coli DH5α, which did not perturb actin assembly appreciably, remained unaffected by rWnt5A treatment. Phagosomes prepared separately from Wnt5A conditioned medium treated K1 and K12-MG1655 infected macrophages revealed differences in the relative levels of actin and actin network promoting proteins, upholding that the Wnt5A-Actin axis operates differently for internalized pathogen and non-pathogen. Interestingly, exposure of rWnt5A treated K1 and K12-MG1655/DH5α infected macrophages to actin assembly inhibitors reversed the scenario, blocking killing of K1, yet promoting killing of both K12-MG1655 and DH5α. Taken together, our study illustrates that the state of activation of the Wnt5A/Actin axis in the context of the incumbent bacteria is crucial for directing host response to infection.

ESSENTIAL OIL ANALYSIS OF THE „MICROMERIA JULIANA“ (L.) BENTH., FROM LUŠTICA, MONTENEGRO

Micromeria (Bentham, 1829.) is a polymorph genus of Lamiaceae with about 130 species grouped in three sections: Cymularia, Eumicromeria and Pseudomelissa, the most diverse in Mediterranean regions, but widespread across Europe, Asia, Africa, and North America [1]. Up to now, eight Micromeria species have been recorded inhabiting the territory of Montenegro including a dwarf shrub, Micromeria juliana (L.) Benth. Knowing the existence of the variability in quantity and qualitative makeup, interpopulational, and the interspecies variability of the up to now reported essential oils from this plant prompted us to make the detailed analysis of the M. juliana specimens originating from rocky, sunny carbonate rock bases in Zabrđe, Montenegro, since the plant material from this area was not examined before. Detailed GC-MS analysis of the obtained essential oil resulted in 70 identified components among which α-pinene (12.2%), (E)-nerolidol (8.9%), viridiflorol (6.8%), limonene (6.1%), and borneol (5.2%) prevailed, differing significantly in content from the previously reported populations (Morača canyon, Cijevna canyon, Mt. Orjen and Mt. Krivošije) [2]. The objective of this study is to gain insight into the essential oil structure, determine the possible mutual similarity and the variability in their chemical compositions, and to enclose possible diversity in the secondary metabolite profile dependent on the site of collection.