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Characterization of the Skin Cultivable Microbiota Composition of the Frog Pelophylax perezi Inhabiting Different Environments.

ABSTRACT: Microorganisms that live in association with amphibian skin can play important roles in protecting their host. Within the scenarios of global change, it is important to understand how environmental disturbances, namely, metal pollution, can affect this microbiota. The aim of this study is to recognize core bacteria in the skin cultivable microbiota of the Perez frog (Pelophylax perezi) that are preserved regardless of the environmental conditions in which the frogs live. The characterization of these isolates revealed characteristics that can support their contributions to the ability of frogs to use metal impacted environments. Frog's skin swabs were collected from P. perezi populations that inhabit a metal-polluted site and three reference (non-metal polluted) sites. Bacterial strains were isolated, identified, and subjected to an acid mine drainage tolerance (AMD) test, collected upstream from a site heavily contaminated with metals, and tested to produce extracellular polymeric substances (exopolysaccharide, EPS). All frog populations had Acinetobacter in their cutaneous cultivable microbiota. Significant growth inhibition was observed in all bacterial isolates exposed to 75% of AMD. EPS production was considered a characteristic of several isolates. The data obtained is a preliminary step but crucial to sustain that the cultivable microbiota is a mechanism for protecting frogs against environmental contamination.

SUBMITTER: Proenca DN

PROVIDER: S-EPMC7967507 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Water frogs of the genus Pelophylax (previous Rana) species have been much studied in Europe for their outstanding reproductive mechanism in which sympatric hybridization between genetically distinct parental species produces diverse genetic forms of viable hybrid animals. The most common hybrid is P. esculentus that carries the genomes of both parental species, P. ridibundus and P. lessonae, but usually transfers the whole genome of only one parent to its offsprings (hybridogenesis). The evolutionary cost of transfer of the intact genome and hence the hemiclonal reproduction is the depletion of heterozygosity in the hybrid populations. Pelophylax esculentus presents an excellent example of the long-term sustained hybridization and hemiclonal reproduction in which the effects of the low genetic diversity are balanced through the novel mutations and periodic recombinations. In this study, we analyzed the mitochondrial (mt) and microsatellites DNA variations in hybrid Pelophylax populations from southern parts of the Pannonian Basin and a north-south transect of the Balkan Peninsula, which are home for a variety of Pelophylax genetic lineages. The mtDNA haplotypes found in this study corresponded to P. ridibundus and P. epeiroticus of the Balkan - Anatolian lineage (ridibundus-bedriagae) and to P. lessonae and a divergent lessonae haplotype of the lessonae lineage. The mtDNA genomes showed considerable intraspecific variation and geographic differentiation. The Balkan wide distributed P. ridibundus was found in all studied populations and its nuclear genome, along with either the lessonae or the endemic epeiroticus genome, in all hybrids. An unexpected finding was that the hybrid populations were invariably heteroplasmic, that is, they contained the mtDNA of both parental species. We discussed the possibility that such extensive heteroplasmy is a result of hybridization and it comes from regular leakage of the paternal mtDNA from a sperm of one species that fertilizes eggs of another. In this case, the mechanisms that protect the egg from heterospecific fertilization and further from the presence of sperm mtDNA could become compromised due to their differences and divergence at both, mitochondrial and nuclear DNA. The heteroplasmy once retained in the fertilized egg could be transmitted by hybrid backcrossing to the progeny and maintained in a population over generations. The role of interspecies and heteroplasmic hybrid animals due to their genomic diversity and better fitness compare to the parental species might be of the special importance in adaptations to miscellaneous and isolated environments at the Balkan Peninsula.

Project description:Besides nutritional components, breast milk contains diverse microbes, which may be involved in colonization of the infant gut. Expressed milk is often stored for few days in the refrigerator. The aim of this study was to determine the abundance, prevalence and diversity of facultative and strict anaerobic bacteria using culture-dependent and -independent methods, and to determine changes in milk microbial and chemical composition during storage. Samples of mature breast milk from 21 women were collected 3-6 months post-partum and were analyzed fresh or after anaerobic storage for 6 days at 4°C. The cultivable bacterial population was analyzed using the most probable number (MPN) method or plate counts and different media. The abundance of major bacterial groups was determined using quantitative PCR and 16S rRNA gene sequencing. Lactose, lactate, short chain fatty acids (SCFA) and human milk oligosaccharides (HMO) were analyzed using chromatography techniques. Highest mean viable cell counts were obtained in yeast casitone fatty acids (YCFA) broth supplied with mucin (log 4.2 ± 1.8 cells/ml) and lactose (log 3.9 ± 1.4 cells/ml), or Columbia broth (log 3.0 ± 0.7 cells/ml). Mean total bacterial counts estimated by qPCR was 5.3 ± 0.6 log cells/ml, with Firmicutes being the most abundant phylum. The most prevalent bacterial groups were Streptococcus spp. (15/19 samples), Enterobacteriaceae (13/19) and Lactobacillus/Lactococcus/Pediococcus group (12/19). While the average total number of bacterial cells did not change significantly during storage, the prevalence of strict anaerobic Bacteroidetes increased threefold, from 3/19 to 9/19, while in 7 samples Clostridium clusters IV or XIVa became detectable after storage. Major HMO were not degraded. Lactate was present in 18/21 samples after storage (2.3-18.0 mM). Butyrate was detected in 15/21 and 18/21 samples before and after storage, respectively, at concentrations ranging from 2.5 to 5.7 mM. We demonstrate enhanced prevalence and/or abundance of viable strict anaerobes from the Bacteroidetes and Clostridiales after 6-day anaerobic storage of human milk. Our data indicate that anaerobic cold storage did not markedly change total viable bacterial load, while HMO profiles were stable. Anaerobic cold storage of human milk for up to 6 days may be suitable for preserving milk quality for potential microbial transfer to the infant gut.

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Characterization of the Skin Cultivable Microbiota Composition of the Frog Pelophylax perezi Inhabiting Different Environments.

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