Bacteriocyte dynamics during development of a holometabolous insect, the carpenter ant Camponotus floridanus

<p>Abstract</p> <p>Background</p> <p>The carpenter ant <it>Camponotus floridanus </it>harbors obligate intracellular mutualistic bacteria (<it>Blochmannia floridanus</it>) in specialized cells, the bacteriocytes, intercalated in their midgut tissue. The diffuse distribution of bacteriocytes over the midgut tissue is in contrast to many other insects carrying endosymbionts in specialized tissues which are often connected to the midgut but form a distinct organ, the bacteriome. <it>C. floridanus </it>is a holometabolous insect which undergoes a complete metamorphosis. During pupal stages a complete restructuring of the inner organs including the digestive tract takes place. So far, nothing was known about maintenance of endosymbionts during this life stage of a holometabolous insect. It was shown previously that the number of <it>Blochmannia </it>increases strongly during metamorphosis. This implicates an important function of <it>Blochmannia </it>in this developmental phase during which the animals are metabolically very active but do not have access to external food resources. Previous experiments have shown a nutritional contribution of the bacteria to host metabolism by production of essential amino acids and urease-mediated nitrogen recycling. In adult hosts the symbiosis appears to degenerate with increasing age of the animals.</p> <p>Results</p> <p>We investigated the distribution and dynamics of endosymbiotic bacteria and bacteriocytes at different stages during development of the animals from larva to imago by confocal laser scanning microscopy. The number of bacteriocytes in relation to symbiont-free midgut cells varied strongly over different developmental stages. Especially during metamorphosis the relative number of bacteria-filled bacteriocytes increased strongly when the larval midgut epithelium is shed. During this developmental stage the midgut itself became a huge symbiotic organ consisting almost exclusively of cells harboring bacteria. In fact, during this phase some bacteria were also found in midgut cells other than bacteriocytes indicating a cell-invasive capacity of <it>Blochmannia</it>. In adult animals the number of bacteriocytes generally decreased.</p> <p>Conclusions</p> <p>During the life cycle of the animals the distribution of bacteriocytes and of <it>Blochmannia </it>endosymbionts is remarkably dynamic. Our data show how the endosymbiont is retained within the midgut tissue during metamorphosis thereby ensuring the maintenance of the intracellular endosymbiosis despite a massive reorganization of the midgut tissue. The transformation of the entire midgut into a symbiotic organ during pupal stages underscores the important role of <it>Blochmannia </it>for its host in particular during metamorphosis.</p

On strongly walk regular graphs, triple sum sets and their codes

Strongly walk-regular graphs (SWRGs) can be constructed as coset graphs of the duals of projective three-weight codes whose weights satisfy a certain equation. We provide classifications of the feasible parameters of these codes in the binary and ternary case for medium size code lengths. For the binary case, the divisibility of the weights of these codes is investigated and several general results are shown. It is known that an SWRG has at most 4 distinct eigenvalues $k > \theta_1 > \theta_2 > \theta_3$. For an $s$-SWRG, the triple $(\theta_1, \theta_2, \theta_3)$ satisfies a certain homogeneous polynomial equation of degree $s - 2$ (Van Dam, Omidi, 2013). This equation defines a plane algebraic curve; we use methods from algorithmic arithmetic geometry to show that for $s = 5$ and $s = 7$, there are only the obvious solutions, and we conjecture this to remain true for all (odd) $s \ge 9$.Comment: 42 page

On strongly walk regular graphs,triple sum sets and their codes

Strongly walk-regular graphs (SWRGs) can be constructed as coset graphs of the duals of projective three-weight codes whose weights satisfy a certain equation. We provide classifications of the feasible parameters of these codes in the binary and ternary case for medium size code lengths. For the binary case, the divisibility of the weights of these codes is investigated and several general results are shown. It is known that an SWRG has at most 4 distinct eigenvalues $k > \theta_1 > \theta_2 > \theta_3$. For an $s$-SWRG, the triple $(\theta_1, \theta_2, \theta_3)$ satisfies a certain homogeneous polynomial equation of degree $s - 2$ (Van Dam, Omidi, 2013). This equation defines a plane algebraic curve; we use methods from algorithmic arithmetic geometry to show that for $s = 5$ and $s = 7$, there are only the obvious solutions, and we conjecture this to remain true for all (odd) $s \ge 9$.Comment: 42 page

Performance and Recovery of Well-Trained Younger and Older Athletes during Different HIIT Protocols

Due to physiological and morphological differences, younger and older athletes may recover differently from training loads. High-intensity interval training (HIIT) protocols are useful for studying the progression of recovery. It was the objective of this study to determine age differences in performance and recovery following different HIIT protocols. Methods: 12 younger (24.5 ± 3.7 years) and 12 older (47.3 ± 8.6 years) well-trained cyclists and triathletes took part in this study. Between the age groups there were no significant differences in relative peak power to fat-free mass, maximal heart rate (HR), training volume, and VO2max-percentiles (%). Participants performed different HIIT protocols consisting of 4 × 30 s Wingate tests with different active rest intervals (1, 3, or 10 min). Peak and average power, lactate, HR, respiratory exchange ratio (RER), subjective rating of perceived exertion (RPE), and recovery (Total Quality Recovery scale, TQR) were assessed. Results: During the different HIIT protocols, metabolic, cardiovascular, and subjective recovery were similar between the two groups. No significant differences were found in average lactate concentration, peak and average power, fatigue (%), %HRmax, RER, RPE, and TQR values between the groups (p > 0.05). Conclusion: The findings of this study indicate that recovery following HIIT does not differ between the two age groups. Furthermore, older and younger participants displayed similar lactate kinetics after the intermittent exercise protocols

Midlatitude shelf seas in the Cenomanian-Turonian greenhouse world: Temperature evolution and North Atlantic circulation

An 8 million year record of subtropical and midlatitude shelf-sea temperatures, derived from oxygen isotopes of well-preserved brachiopods from a variety of European sections, demonstrates a long-term Cenomanian temperature rise (16–20°C, midlatitudes) that reached its maximum early in the late Turonian (23°C, midlatitudes). Superimposed on the long-term trend, shelf-sea temperatures vary at shorter timescales in relation to global carbon cycle perturbations. In the mid-Cenomanian and the late Turonian, two minor shelf-sea cooling events (2–3°C) coincide with carbon cycle perturbations and times of high-amplitude sea level falls. Although this evidence supports the hypothesis of potential glacioeustatic effects on Cretaceous sea level, the occurrence of minimum shelf-sea temperatures within transgressive beds argues for regional changes in shelf-sea circulation as the most plausible mechanism. The major carbon cycle event in the latest Cenomanian (oceanic anoxic event 2) is accompanied by a substantial increase in shelf-sea temperatures (4–5°C) that occurred ∼150 kyr after the commencement of the δ13C excursion and is related to the spread of oceanic conditions in western European shelf-sea basins. Our oxygen isotope record and published δ18O data of pristinely preserved foraminifera allow the consideration of North Atlantic surface water properties in the Cenomanian along a transect from the tropics to the midlatitudes. On the basis of fossil-derived δ18O, estimated δw ranges, and modeled salinities, temperature-salinity-density ranges were estimated for tropical, subtropical, and midlatitude surface waters. Accordingly, the Cenomanian temperate shelf-seas waters have potentially the highest surface water density and could have contributed to North Atlantic intermediate to deep waters in the preopening stage of the equatorial Atlantic gateway

Nutritional upgrading for omnivorous carpenter ants by the endosymbiont Blochmannia

<p>Abstract</p> <p>Background</p> <p>Carpenter ants (genus <it>Camponotus</it>) are considered to be omnivores. Nonetheless, the genome sequence of <it>Blochmannia floridanus</it>, the obligate intracellular endosymbiont of <it>Camponotus floridanus</it>, suggests a function in nutritional upgrading of host resources by the bacterium. Thus, the strongly reduced genome of the endosymbiont retains genes for all subunits of a functional urease, as well as those for biosynthetic pathways for all but one (arginine) of the amino acids essential to the host.</p> <p>Results</p> <p>Nutritional upgrading by <it>Blochmannia </it>was tested in 90-day feeding experiments with brood-raising in worker-groups on chemically defined diets with and without essential amino acids and treated or not with antibiotics. Control groups were fed with cockroaches, honey water and Bhatkar agar. Worker-groups were provided with brood collected from the queenright mother-colonies (45 eggs and 45 first instar larvae each). Brood production did not differ significantly between groups of symbiotic workers on diets with and without essential amino acids. However, aposymbiotic worker groups raised significantly less brood on a diet lacking essential amino acids. Reduced brood production by aposymbiotic workers was compensated when those groups were provided with essential amino acids in their diet. Decrease of endosymbionts due to treatment with antibiotic was monitored by qRT-PCR and FISH after the 90-day experimental period. Urease function was confirmed by feeding experiments using ¹⁵N-labelled urea. GC-MS analysis of ¹⁵N-enrichment of free amino acids in workers revealed significant labelling of the non-essential amino acids alanine, glycine, aspartic acid, and glutamic acid, as well as of the essential amino acids methionine and phenylalanine.</p> <p>Conclusion</p> <p>Our results show that endosymbiotic <it>Blochmannia </it>nutritionally upgrade the diet of <it>C. floridanus </it>hosts to provide essential amino acids, and that it may also play a role in nitrogen recycling via its functional urease. <it>Blochmannia </it>may confer a significant fitness advantage via nutritional upgrading by enhancing competitive ability of <it>Camponotus </it>with other ant species lacking such an endosymbiont. Domestication of the endosymbiont may have facilitated the evolutionary success of the genus <it>Camponotus</it>.</p

Scholte-wave tomography for shallow-water marine sediments

We determine the 3-D in situ shear-wave velocities of shallow-water marine sediments by extending the method of surface wave tomography to Scholte-wave records acquired in shallow waters. Scholte waves are excited by air-gun shots in the water column and recorded at the seafloor by ocean-bottom seismometers as well as buried geophones. Our new method comprises three steps: (1) We determine local phase-slowness values from slowness-frequency spectra calculated by a local wavefield transformation of common-receiver gathers. Areal phase-slowness maps for each frequency used as reference in the following step are obtained by interpolating the values derived from the local spectra. (2) We infer slowness residuals to those reference slowness maps by a tomographic inversion of the phase traveltimes of fundamental Scholte-wave mode. (3) The phase-slowness maps together with the residuals at different frequencies define a local dispersion curve at every location of the investigation area. From those dispersion curves we determine a model of the depth-dependency of shear-wave velocities for every location. We apply this method to a 1 km2 investigation area in the Baltic Sea (northern Germany). The phase-slowness maps obtained in step (2) show lateral variation of up to 150 per cent. The shear-wave velocity models derived in the third step typically have very low values (60–80 m s−1) in the top four meters where fine muddy sands can be observed, and values exceeding 170 m s−1 for the silts and sands below that level. The upper edge of glacial till with shear-wave velocities of 300–400 m s−1 is situated approximately 20 m below sea bottom. A sensitivity analysis reveals a maximum penetration depth of about 40 m below sea bottom, and that density may be an important parameter, best resolvable with multimode inversion

Functional analysis of Blochmannia floridanus, the primary endosymbiont of the carpenter ant Camponotus floridanus

Ameisen der Gattung Camponotus beherbergen bakterielle Symbionten der Gattung Blochmannia in spezialisierten Zellen des Mitteldarms (Blochmann, 1882; Buchner, 1965; Sauer, 2000; Schröder et al., 1996). Die Genomsequenzierung dieser Symbionten zeigte, dass Blochmannia, ähnlich den Symbionten von Blattläusen, hauptsächlich Gene der Aminosäurebiosynthese beibehalten hat (Degnan et al., 2005; Gil et al., 2003). Die Relevanz dieser nahrungsaufwertenden Funktion konnte experimentell bestätigt werden (Feldhaar et al., 2007). Ein Schwerpunkt der vorliegenden Arbeit war die Aufklärung der dynamischen Interaktion der beiden Partner während des komplexen Lebenszyklus des holometabolen Wirtes. Frühere Studien deuteten darauf hin, dass die Symbiose vor allem während der Larven- und Puppenphasen von Bedeutung sein könnte (Feldhaar et al., 2007; Wolschin et al., 2004; Zientz et al., 2006). Mit fluoreszenter in situ Hybridisierung (FISH) und konfokaler Laserscanning Mikroskopie konnte in der vorliegenden Arbeit die Lokalisierung von B. floridanus während der wichtigsten Entwicklungsstadien aufgeklärt werden. Hierbei konnte gezeigt werden, dass die Symbionten schon im ersten Larvenstadium in spezialisierten Zellen um den Darm angeordnet sind, aber in späteren Stadien nicht, wie bisher angenommen, auf diese Bakteriozyten beschränkt sind, sondern bis zum Schlupf der jungen Arbeiterinnen massiv andere Darmzellen infizieren. Übereinstimmend mit Bestimmungen der Zellzahl in den verschiedenen Wirtsstadien ist die Anzahl der Symbionten gegen Ende der Metamorphose am höchsten. Die Symbiose degeneriert in sehr alten Arbeiterinnen, gut gefüllte Bakteriozyten werden jedoch noch monatelang beibehalten. Mit Macroarray- und qRT- PCR- basierten Transkriptomanalysen wurde die Expression der bakteriellen Gene in charakteristischen Entwicklungsstadien des Wirtes untersucht. Allgemein zeigen vor allem Gene für molekulare Chaperons und bestimmte bakterielle Grundfunktionen eine hohe Expression. Aber auch viele Gene, die möglicherweise wichtige Funktionen in der Symbiose besitzen, wie die Biosynthese essentieller Aminosäuren und das Recycling von Stickstoffverbindungen, zeigen ein hohes absolutes Transkriptlevel. Zudem besteht eine positive Korrelation zwischen dem Expressionsniveau und dem GC- Gehalt der Gene, die in dem höheren Selektionsdruck und damit einer geringeren Mutationsrate der essentiellen Gene begründet liegt (Schaber et al., 2005). Durch Proteinanalysen konnte bestätigt werden, dass die Faktoren mit der höchsten absoluten Transkription die dominanten Proteine der Symbionten darstellen. In den unterschiedlichen Entwicklungsstadien zeigen viele Gene eine deutliche Dynamik, deren Ausmaß aber, verglichen mit freilebenden Bakterien, gering ist. Aus den Expressionsprofilen aufeinanderfolgender Gene lassen sich mögliche Transkriptionseinheiten ableiten, die teilweise auch experimentell bestätigt wurden. Oftmals zeigen auch Gene, die nicht in Transkriptionseinheiten angeordnet sind, aber verwandten Stoffwechselwegen angehören, ähnliche Muster. Dies deutet auf das Vorhandensein grundlegender Genregulations-mechanismen hin, obwohl im Genom von B. floridanus nur noch sehr wenige Transkriptionsfaktoren codiert sind (Gil et al., 2003). Auf übergeordneter Ebene zeigt sich, dass bei Symbionten aus späten Puppenstadien viele symbioserelevante Gene im Vergleich zu Genen des Grundmetabolismus eine erhöhte Expression zeigen. Dies betrifft besonders die Biosynthese aromatischer und verzweigter Aminosäuren, die in diesen Stadien vom Wirt in hoher Menge benötigt werden, während die internen Reserven gleichzeitig zur Neige gehen. Dies äußert sich auch im deutlichen Abfallen der Speicherproteinmenge des Wirts gegen Ende der Puppenphase. Die festgestellte Veränderung der Symbiontenzahl übertrifft das geringe Ausmaß der Genregulation um ein Vielfaches. Die Bakterien liegen in jedem Stadium polyploid mit bis zu 100 Genomkopien vor, dieser Polyploidiegrad bleibt jedoch während der gesamten Wirtsentwicklung weitestgehend konstant. Somit scheint die Kontrolle des Wirts über die bakterielle Vermehrung der entscheidende Faktor dieser Symbiose zu sein. Die verbleibenden regulatorischen Fähigkeiten der Bakterien stellen möglicherweise eine Feinjustierung von optimierten Produktionseinheiten dar, deren Anzahl nach den Bedürfnissen des Wirtes verändert wird. Insgesamt konnten in der vorliegenden Arbeit neue Einblicke in das komplexe Zusammenleben von Blochmannia und Camponotus gewonnen werden, die zu einem besseren Verständnis der biologischen Funktion und der grundlegenden Mechanismen dieser Symbiose führen. Eine der wichtigsten Fragestellungen nach dem Sinn einer nahrungsaufwertenden Symbiose für einen Nahrungsgeneralisten konnte mit starken Hinweisen auf eine stadienabhängige Relevanz der Symbiose beantwortet werden, die den enormen evolutionären Erfolg dieser Ameisengattung erklären könnte.&#8195;Ants of the genus Camponotus harbor bacterial endosymbionts of the genus Blochmannia in specialized cells of their midgut (Blochmann, 1882; Buchner, 1965; Sauer, 2000; Schröder et al., 1996). The complete sequencing of the symbiont’s genome revealed, that Blochmannia, comparable to the symbionts of aphids, mainly retained genes involved in the biosynthesis of essential amino acids (Degnan et al., 2005; Gil et al., 2003). The biological relevance of a nutritional upgrading by Blochmannia could be confirmed experimentally (Feldhaar et al., 2007). One focus of this thesis was the elucidation of the dynamic interactions between the two partners during the complex life cycle of the holometabolic host animal. Previous studies pointed towards a temporal relevance of this symbiosis especially during larval and pupal development (Feldhaar et al., 2007; Wolschin et al., 2004; Zientz et al., 2006). In this thesis the localization of B. floridanus could be documented throughout all life stages of the host by fluorescent in situ hybridization (FISH) and confocal laser scanning microscopy. A layer of densely filled bacteriocytes surrounding the gut could already be identified in first instar larvae. In contrast to previous assumptions, the bacteria are not restricted to these cells in later stages, as until the eclosion of the young adult workers bacteria massively infect other midgut cells. Concordant with previous findings, bacterial load is highest at the end of metamorphosis and symbiont numbers decrease in older workers, yet densely filled bacteriocytes are still visible after several months. The expression of the bacterial genes during characteristic life stages of the C. floridanus was assessed by macroarray and qRT- PCR- based experiments. In general, especially molecular chaperones, central basic metabolism and may putative symbiosis related factors like pathways leading to essential amino acids or nitrogen recycling show highest absolute expression levels. A positive correlation between expression level and GC- content of the genes can be observed, which is caused by a higher selection pressure and lower mutation rate of these essential factors (Schaber et al., 2005). Protein analyses confirmed the correlation between gene expression and translation of the most abundant factors. Many B. floridanus genes exhibit a dynamic expression during the different host stages but the extent of this gene regulation is modest as compared to free living bacteria. Expression profiles of genes located next to each other on the genome allow proposal of local transcription units, which were confirmed experimentally in several cases. Often genes that are not clustered locally but belong to related metabolic functions also exhibit similar expression patterns. This indicates the existence of basic mechanisms of gene regulation despite the low number of transcription factors annotated in the B. floridanus genome (Gil et al., 2003). In late pupal stages symbiosis related genes often show a higher expression compared to basic metabolic functions. This especially includes biosynthetic pathways for aromatic and branched amino acids, which are needed by the host at this stage in increased amounts, while internal storages are depleted. This could be demonstrated by the significant decrease in storage proteins of the host at the end of the pupal phase. The observed change in bacterial numbers per host exceeds the extent of bacterial gene regulation by far. The symbionts are polyploid in each host stage with up to 100 genome copies per cell. The degree of polyploidy is largely constant during host development. Thus the control over bacterial reproduction seems to be the decisive factor in this symbiosis. The residual regulatory capacities of the symbionts might represent a mechanism of fine tuning of a production unit that has been streamlined by evolution and whose numbers are adjusted according to the host’s needs. In conclusion, this thesis delivers new insights into the complex symbiosis of Blochmannia and Camponotus leading to a better understanding of its biological function and the underlying mechanisms. One of the central mysteries concerning the need of a symbiont for nutritional upgrading for an omnivorous host could be explained by a temporal, stage- dependent relevance of this symbiosis, possibly being the reason for the enormous evolutionary success of this ant genus

On strongly walk regular graphs, triple sum sets and their codes

International audienceStrongly walk regular graphs (SWRGs or s-SWRGs) form a natural generalization of strongly regular graphs (SRGs) where paths of length 2 are replaced by paths of length s. They can be constructed as coset graphs of the duals of projective three-weight codes whose weights satisfy a certain equation. We provide classifications of the feasible parameters of these codes in the binary and ternary case for medium size code lengths. For the binary case, the divisibility of the weights of these codes is investigated and several general results are shown. It is known that an s-SWRG has at most 4 distinct eigenvalues k > θ 1 > θ 2 > θ 3 , and that the triple (θ 1 , θ 2 , θ 3) satisfies a certain homogeneous polynomial equation of degree s − 2 (Van Dam, Omidi, 2013). This equation defines a plane algebraic curve; we use methods from algorithmic arithmetic geometry to show that for s = 5 and s = 7, there are only the obvious solutions, and we conjecture this to remain true for all (odd) s ≥ 9