mtDNA diversity in rhesus monkeys reveals overestimates of divergence time and paraphyly with neighboring species

Reconstructions of the human-African great ape phylogeny by using mitochondrial DNA (mtDNA) have been subject to considerable debate. One confounding factor may be the lack of data on intraspecific variation. To test this hypothesis, we examined the effect of intraspecific mtDNA diversity on the phylogenetic reconstruction of another Plio- Pleistocene radiation of higher primates, the fascicularis group of macaque (Macaca) monkey species. Fifteen endonucleases were used to identify 10 haplotypes of 40-47 restriction sites in M. mulatta, which were compared with similar data for the other members of this species group. Interpopulational, intraspecific mtDNA diversity was large (0.5%- 4.5%), and estimates of divergence time and branching order incorporating this variation were substantially different from those based on single representatives of each species. We conclude that intraspecific mtDNA diversity is substantial in at least some primate species. Consequently, without prior information on the extent of genetic diversity within a particular species, intraspecific variation must be assessed and accounted for when reconstructing primate phylogenies. Further, we question the reliability of hominoid mtDNA phylogenies, based as they are on one or a few representatives of each species, in an already depauperate superfamily of primates.      

Culture of bovine embryos after invitro exposure to Brucellaabortus

Fifty-four day-6 through day-10 (estrus=day 0) embryos were collected nonsurgically from 13 superovulated, brucellosis-free mixed breed cows. Forty-eight excellent and good zona pellucida-intact (ZP-I), three zona pellucida-defective (ZP-D), and three zona pellucida-free (ZP-F) embryos were incubated in media containing $\text{Brucella}$$\text{abortus}$. Subsequently, embryos were washed ten times in groups of one, two, three, or four. Embryos and serial washes were cultured for $\text{B}$. $\text{abortus}$.Brucellae were not isolated from any ZP-I embryo or from any washing beyond the sixth serial wash. Brucellae were not isolated from the three ZP-F embryos but were detected in the eighth wash for one and in the tenth wash for the others. Brucellae were isolated from one of three ZP-D embryos. Results show that ZP-I embryos can be effectively washed free of $\text{B}$. $\text{abortus}$.     

Lipid remodelling is a widespread strategy in marine heterotrophic bacteria upon phosphorus deficiency

Upon phosphorus (P) deficiency, marine phytoplankton reduce their requirements for P by replacing membrane phospholipids with alternative non-phosphorus lipids. It was very recently demonstrated that a SAR11 isolate also shares this capability when phosphate starved in culture. Yet, the extent to which this process occurs in other marine heterotrophic bacteria and in the natural environment is unknown. Here, we demonstrate that the substitution of membrane phospholipids for a variety of non-phosphorus lipids is a conserved response to P deficiency among phylogenetically diverse marine heterotrophic bacteria, including members of the Alphaproteobacteria and Flavobacteria. By deletion mutagenesis and complementation in the model marine bacterium Phaeobacter sp. MED193 and heterologous expression in recombinant Escherichia coli, we confirm the roles of a phospholipase C (PlcP) and a glycosyltransferase in lipid remodelling. Analyses of the Global Ocean Sampling and Tara Oceans metagenome data sets demonstrate that PlcP is particularly abundant in areas characterized by low phosphate concentrations. Furthermore, we show that lipid remodelling occurs seasonally and responds to changing nutrient conditions in natural microbial communities from the Mediterranean Sea. Together, our results point to the key role of lipid substitution as an adaptive strategy enabling heterotrophic bacteria to thrive in the vast P-depleted areas of the ocean.     

2-Aminoethylphosphonate utilization in Pseudomonas putida BIRD-1 is controlled by multiple master regulators

Bacteria possess various regulatory mechanisms to detect and coordinate a response to elemental nutrient limitation. In pseudomonads, the two-component system regulators CbrAB, NtrBC and PhoBR, are responsible for regulating cellular response to carbon (C), nitrogen (N) and phosphorus (P) respectively. Phosphonates are reduced organophosphorus compounds produced by a broad range of biota and typified by a direct C-P bond. Numerous pseudomonads can use the environmentally abundant phosphonate species 2-aminoethylphosphonate (2AEP) as a source of C, N, or P, but only PhoBR has been shown to play a role in 2AEP utilization. On the other hand, utilization of 2AEP as a C and N source is considered substrate inducible. Here, using the plant-growth-promoting rhizobacterium Pseudomonas putida BIRD-1 we present evidence that 2AEP utilization is under dual regulation and only occurs upon depletion of C, N, or P, controlled by CbrAB, NtrBC, or PhoBR respectively. However, the presence of 2AEP was necessary for full gene expression, i.e. expression was substrate inducible. Mutation of a LysR-type regulator, termed AepR, upstream of the 2AEP transaminase-phosphonatase system (PhnWX), confirmed this dual regulatory mechanism. To our knowledge, this is the first study identifying coordination between global stress response and substrate-specific regulators in phosphonate metabolism.     

PCR analysis of the distribution of unicellular cyanobacterial diazotrophs in the Arabian Sea

An oligonucleotide primer, NITRO821R, targeting the 16S rRNA gene of unicellular cyanobacterial N2 fixers was developed based on newly derived sequences from Crocosphaera sp. strain WH 8501 and Cyanothece sp. strains WH 8902 and WH 8904 as well as several previously described sequences of Cyanothece sp. and sequences of intracellular cyanobacterial symbionts of the marine diatom Climacodium frauenfeldianum. This oligonucleotide is specific for the targeted organisms, which represent a well-defined phylogenetic lineage, and can detect as few as 50 cells in a standard PCR when it is used as a reverse primer together with the cyanobacterium- and plastid-specific forward primer CYA359F (U. Nubel, F. Garcia-Pichel, and G. Muyzer, Appl. Environ. Microbiol. 63:3327-3332, 1997). Use of this primer pair in the PCR allowed analysis of the distribution of marine unicellular cyanobacterial diazotrophs along a transect following the 67 degrees E meridian from Victoria, Seychelles, to Muscat, Oman (0.5 degrees S to 26 degrees N) in the Arabian Sea. These organisms were found to be preferentially located in warm (>29 degrees C) oligotrophic subsurface waters between 0 and 7 degrees N, but they were also found at a station north of Oman at 26 degrees N, 56 degrees 35'E, where similar water column conditions prevailed. Slightly cooler oligotrophic waters (<29 degrees C) did not contain these organisms or the numbers were considerably reduced, suggesting that temperature is a key factor in dictating the abundance of this unicellular cyanobacterial diazotroph lineage in marine environments.     

Real-Time, label-free monitoring of tumor antigen and serum antibody interactions

Conventional techniques for the detection of biomolecular interactions can be limited by the need for exogenous labels, time- and labor-intensive protocols, as well as by poor sensitivity levels. A refractometer instrument has been reconfigured to detect biomolecular interactions through changes in surface plasmon resonance (SPR). The binding kinetics and affinity values of anti-NY-ESO-1 monoclonal antibody, ES121, to the cancer-testis antigen NY-ESO-1 were determined according to the surface heterogeneity model and resulted in K(D) values of 1.3x10(-9) and 2.1x10(-10) M. The reconfigured instrument was then used to measure the interaction between tumor antigens and serum antibodies against these antigens in preselected cancer patient sera samples. The tumor antigens assayed included NY-ESO-1, SSX2 and p53, all used as recombinant proteins containing polyhistidine tags. These results demonstrated that the instrument is capable of detecting the binding of serum antibodies from cancer patient sera to immobilized tumor antigens, consistent with those observed previously in ELISA-based experiments. These results demonstrate the potential of SPR technology for the rapid diagnosis and monitoring immune responses.     

Estradiol modulation of growth hormone secretion in the ewe: no growth hormone-releasing hormone neurons and few somatotropes express estradiol receptor alpha

Evidence suggests that estrogen modulates growth hormone (GH) release and that GH plays an important role in follicular and ovulatory processes. How estradiol affects GH secretion is unclear. Having verified that there is a coincident surge of GH at the time of the preovulatory LH surge, immunocytochemical studies incorporating high-temperature antigen retrieval were used to determine whether GH-releasing hormone (GHRH) neurons, somatotropes, or both, expressed estrogen receptor alpha (ER), in the ewe. Although GHRH neurons were surrounded by many ER cells, they did not express immunocytochemically detectable ERs. In contrast to gonadotropes, in which the majority expressed ERs, few somatotropes were estrogen receptive. These data suggest that estrogen does not act directly on GHRH neurons to influence GH secretion, and any direct effect on pituitary GH release, through the ERalpha, may be small.