Characterization of the gut microbiome and resistome of Galapagos marine iguanas (Amblyrhynchus cristatus) from uninhabited islands.

BACKGROUND: Understanding the natural microbiome and resistome of wildlife from remote places is necessary to monitor the human footprint on the environment including antimicrobial use (AU). Marine iguanas are endemic species from the Galapagos Islands where they are highly affected by anthropogenic factors that can alter their microbiota as well as their abundance and diversity of antimicrobial-resistant genes (ARGs). Thus, this study aims to apply culture-independent approaches to characterize the marine iguana's gut metagenomic composition of samples collected from the uninhabited islands Rabida (n = 8) and Fernandina (Cabo Douglas, n = 30; Punta Espinoza, n = 30). Fresh feces from marine iguanas were analyzed through SmartChip RT-PCR, 16S rRNA, and metagenomic next-generation sequencing (mNGS) to identify their microbiome, microbial-metabolic pathways, resistome, mobilome, and virulome. RESULTS: The marine iguana's gut microbiome composition was highly conserved despite differences in ecological niches, where 86% of taxa were shared in the three locations. However, site-specific differences were mainly identified in resistome, mobilome, virulorome, and metabolic pathway composition, highlighting the existence of factors that induce microbial adaptations in each location. Functional gut microbiome analyses revealed its role in the biosynthesis and degradation of vitamins, cofactors, proteinogenic amino acids, carbohydrates, nucleosides and nucleotides, fatty acids, lipids, and other compounds necessary for the marine iguanas. The overall bacterial ARG abundance was relatively low (0.006%); nevertheless, the presence of genes encoding resistance to 22 drug classes was identified in the iguana's gut metagenome. ARG-carrying contig and co-occurrence network analyses revealed that commensal bacteria are the main hosts of ARGs. Taxa of public health interest such as Salmonella, Vibrio, and Klebsiella also carried multidrug-resistance genes associated with MGEs which can influence the dissemination of ARGs through horizontal gene transfer. CONCLUSION: Marine iguanas depend on the gut microbiome for the biosynthesis and degradation of several compounds through a symbiotic relationship. Niche-specific adaptations were evidenced in the pool of microbial accessory genes (i.e., ARGs, MGEs, and virulence) and metabolic pathways, but not in the microbiome composition. Culture-independent approaches outlined the presence of a diverse resistome composition in the Galapagos marine iguanas from remote islands. The presence of AR pathogens in marine iguanas raises concerns about the dispersion of microbial-resistant threats in pristine areas, highlighting wildlife as sentinel species to identify the impact of AU.

PUP MORTALITY AND EVIDENCE FOR PATHOGEN EXPOSURE IN GALAPAGOS SEA LIONS (ZALOPHUS WOLLEBAEKI) ON SAN CRISTOBAL ISLAND, GALAPAGOS, ECUADOR.

The Galapagos sea lion ( Zalophus wollebaeki ), an endangered species, experiences high pup mortality (up to 100%) in years when El Niño events reduce food supply in the Galapagos Islands. Mortality of pups in non-El Niño years is estimated to be 5% in undisturbed colonies. From 2009 to 2012 we observed high pup mortality (up to 67%) in colonies close to the Galapagos capital, Puerto Baquerizo Moreno, where contact with humans, domestic animals, and rats is frequent. Gross postmortem findings from 54 pups included hemorrhagic lesions in liver and congestion in lungs; histopathology suggested a possible association with infectious diseases. Evidence of Leptospira infection was found in five out of seven samples collected in 2010. Canine distemper viral (CDV) RNA was detected in tissues from six sea lions (in 2011-12), four of which were confirmed by nucleotide sequencing. The absence of CDV antibodies in 109 juvenile animals tested in 2014 at urban and remote colonies could indicate that the CDV infection observed in 2011 was likely confined to a few animals. Our results indicated that Galapagos sea lions have been exposed at least to two pathogens, Leptospira and CDV; however, the impact of these infections on the sea lions is unclear.

Blood gases, biochemistry, and hematology of Galapagos green turtles (Chelonia mydas).

The green turtle, Chelonia mydas, is an endangered marine chelonian with a circum-global distribution. Reference blood parameter intervals have been published for some chelonian species, but baseline hematology, biochemical, and blood gas values are lacking from the Galapagos sea turtles. Analyses were done on blood samples drawn from 28 green turtles captured in two foraging locations on San Cristóbal Island (14 from each site). Of these turtles, 20 were immature and of unknown sex; the other eight were males (five mature, three immature). A portable blood analyzer (iSTAT) was used to obtain near immediate field results for pH, lactate, pO2, pCO2, HCO3-, Hct, Hb, Na, K, iCa, and Glu. Parameter values affected by temperature were corrected in two ways: (1) with standard formulas; and (2) with auto-corrections made by the iSTAT. The two methods yielded clinically equivalent results. Standard laboratory hematology techniques were employed for the red and white blood cell counts and the hematocrit determination, which was also compared to the hematocrit values generated by the iSTAT. Of all blood analytes, only lactate concentrations were positively correlated with body size. All other values showed no significant difference between the two sample locations nor were they correlated with body size or internal temperature. For hematocrit count, the iSTAT blood analyzer yielded results indistinguishable from those obtained with high-speed centrifugation. The values reported in this study provide baseline data that may be useful in comparisons among populations and in detecting changes in health status among Galapagos sea turtles. The findings might also be helpful in future efforts to demonstrate associations between specific biochemical parameters and disease.