Bioinformatic Characterization and Molecular Evolution of the Lucina pectinata Hemoglobins.

(1) Introduction: Lucina pectinata is a clam found in sulfide-rich mud environments that has three hemoglobins believed to be responsible for the transport of hydrogen sulfide (HbILp) and oxygen (HbIILp and HbIIILp) to chemoautotrophic endosymbionts. The physiological roles and evolution of these globins in sulfide-rich environments are not well understood. (2) Methods: We performed bioinformatic and phylogenetic analyses with 32 homologous mollusk globin sequences. Phylogenetics suggests a first gene duplication resulting in sulfide binding and oxygen binding genes. A more recent gene duplication gave rise to the two oxygen-binding hemoglobins. Multidimensional scaling analysis of the sequence space shows evolutionary drift of HbIILp and HbIIILp, while HbILp was closer to the Calyptogena hemoglobins. Further corroboration is seen by conservation in the coding region of hemoglobins from L. pectinata compared to those from Calyptogena. (3) Conclusions: Presence of glutamine in position E7 in organisms living in sulfide-rich environments can be considered an adaptation to prevent loss of protein function. In HbILp a substitution of phenylalanine in position B10 is accountable for its unique reactivity towards H2S. It appears that HbILp has been changing over time, apparently not subject to functional constraints of binding oxygen, and acquired a unique function for a specialized environment.

Characterization of Recombinant His-Tag Protein Immobilized onto Functionalized Gold Nanoparticles.

The recombinant polyhistidine-tagged hemoglobin I ((His)6-rHbI) from the bivalve Lucina pectinata is an ideal biocomponent for a hydrogen sulfide (H2S) biosensor due to its high affinity for H2S. In this work, we immobilized (His)6-rHbI over a surface modified with gold nanoparticles functionalized with 3-mercaptopropionic acid complexed with nickel ion. The attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) analysis of the modified-gold electrode displays amide I and amide II bands characteristic of a primarily α-helix structure verifying the presence of (His)6-rHbI on the electrode surface. Also, X-ray photoelectron spectroscopy (XPS) results show a new peak after protein interaction corresponding to nitrogen and a calculated overlayer thickness of 5.3 nm. The functionality of the immobilized hemoprotein was established by direct current potential amperometry, using H2S as the analyte, validating its activity after immobilization. The current response to H2S concentrations was monitored over time giving a linear relationship from 30 to 700 nM with a corresponding sensitivity of 3.22 × 10-3 nA/nM. These results confirm that the analyzed gold nanostructured platform provides an efficient and strong link for polyhistidine-tag protein immobilization over gold and glassy carbon surfaces for a future biosensors development.

Reversible Detection and Quantification of Hydrogen Sulfide by Fluorescence Using the Hemoglobin I from Lucina pectinata.

A new detection system for the endogenous gaseous transmitter and environmental pollutant hydrogen sulfide is presented. It is based on the modulation of the fluorescence spectrum of a coumarin dye by the absorption spectrum of the recombinant hemoglobin I from clam Lucina pectinata upon coordination of the analyte. While we establish that the reported affinity of rHbI for H2S has been overestimated, the association of the protein with an appropriate fluorophore allows fast, easy, and reversible detection and quantification of hydrogen sulfide in buffer as well as biological fluids such as human plasma, with a quantification limit around 200 nM at pH 7.4.

Parasites of economically important bivalves from the southern coast of Bahia State, Brazil / Parasitos de bivalves de interesse econômico no Litoral Sul do Estado da Bahia, Brasil

This study investigated the parasites of three commercially important bivalve species (Crassostrea rhizophorae, Mytella guyanensis and Lucina pectinata) from the southern coast of Bahia, Brazil. A total of 540 specimens were collected in August 2009 and February 2010, at three localities. The bivalve specimens were measured on their longest axis, opened, and macroscopically examined for the presence of parasites or signs of disease. They were then fixed in Davidson' solution and subjected to routine histological processing, with paraffin embedding and H&E staining; next, the specimens were examined under a light microscope. No parasites were observed associated with L. pectinata. Rickettsia-like organisms (RLOs), Sphenophrya sp. (Ciliophora), Nematopsis sp. (Apicomplexa), Urastoma sp. (Turbellaria) and Bucephalus sp. (Digenea) were observed in both C. rhizophorae and M. guyanensis, as well as Ancistrocoma sp. (Ciliophora) and Tylocephalum sp. (Cestoda) in the former. A high prevalence of Nematopsis sp. was seen, but caused no apparent damage to the host. Bucephalus sp. caused the destruction of tissues, with castration, but showed low prevalence. The other parasites occurred in low prevalence and intensity, without causing significant damage.(AU)

Neste estudo foram investigados os parasitos de três espécies de bivalves de interesse econômico (Crassostrea rhizophorae, Mytella guyanensis e Lucina pectinata) da Bahia. Foram analisados 540 exemplares, obtidos em duas coletas (agosto-2009 e fevereiro-2010), em três localidades. Os bivalves foram medidos quanto ao seu maior eixo, abertos e examinados macroscopicamente quanto à presença de parasitos ou sinais de enfermidades. Depois disso, foram fixados em solução de Davidson e processados por rotina de histologia, com inclusão em parafina e coloração com H&E. O material foi examinado ao microscópio de luz. Nenhum parasito esteve associado a L. pectinata. Bactérias do tipo RLOs (organismos assemelhados a Rickettsia), Sphenophrya sp. (Ciliophora), Nematopsis sp. (Apicomplexa), Urastoma sp. (Turbellaria) e Bucephalus sp. (Digenea) foram observados tanto em C. rhizophorae quanto em M. guyanensis, sendo que a primeira apresentou ainda Ancistrocoma sp. (Ciliophora) e Tylocephalum sp. (Cestoda). Nematopsis sp. ocorreu em alta prevalência, porém, aparentemente, não causou danos aos bivalves. Bucephalus sp. causou destruição de tecidos, com castração, mas foi pouco prevalente. Os demais parasitos ocorreram em baixa prevalência e intensidade de infecção e sem causar danos.(AU)

Ultrastructural studies of spermiogenesis of the clamLucina pectinata (Bivalvia, Lucinidae) from Northern Brazil

Spermatogenesis and sperm ultrastructure of the clam Lucina pectinata (Lucinidae) from northern Brazil were studied using light and transmission electron microscopy. The uniflagellate spermatozoa are grouped into characteristic rings within somatic cells (Sertoli-like cells), with the acrosome oriented toward the peripheryof these cells. The spermatozoa are long cells of the primitive type (ect-aquasperm) with a total length of 50.2 ± 2.5 mm, consisting of head (acrosome + nucleus), midpiece and tail. Acrosome is formed by an, acrosomal vesicle with a conical cylinder-like shaped (0.9 ± 0.1 mm length and 0.4 ± 0.1 mm in basal diameter) having a deeply infolded basis occupied by the subacrosomal space, containing flocculent material without axial rod. The acrosomal vesicle is formed by a membrane-bounded containing a broad basal ring of electron-dense material. The nucleus (7.5 ± 0.8 mm long) is an elongated, subcylindrical rod-shaped, slightly and gently curved, with a basal invagination (0.2-0.4 mm). It contains dense chromatin without any electron-lucent lacunae.The midpiece (1.0 ± 0.3 mm long; 1.1 ± 0.2 mm width) consist of four equal mitochondria located at the same level, surrounding two centrioles arranged at right angles. The proximal centriole lies at 90° relative the distal centriole and sperm longitudinal axis. The tail (40.5 ± 2.1 mm long) contains the common 9 + 2 pattern which in tapering end piece is successively reduced and sheated by the plasmalemma.