Macroinvertebrate communities associated with duckweed (Lemnaceae) in two Eastern Cape rivers,South Africa

The functional feeding groups and diversity of macroinvertebrate communities associated with duckweed mats in the New Years River (two sites) and Bloukrans River (two sites), Eastern Cape province, South Africa, were assessed. Duckweed (Lemnaceae) is a ubiquitous family of floating macrophytes. A total of 41 macroinvertebrate families were collected monthly over a six-month period from February to July 2014. Duckweed biomass in both rivers was highly variable both temporally and spatially. The majority of identified macroinvertebrate taxa were predators and detritivores, with a small percentage of herbivores. An average of approximately 26% of the macroinvertebrate taxa found were from families that include species from more than one functional feeding group. Although overall measures of diversity and ecosystem health (Fisher’s α and Simpson’s index) remained constant over time in the New Years River, significant differences in macroinvertebrate community structure were seen between sites and months on both rivers, with dissimilarity being driven by a larger number of species in the New Years River. This high variability within macroinvertebrate assemblages probably reflects a combination of heterogeneous duckweed distribution, variation in physico-chemistry, opportunistic behaviours of macroinvertebrate predators and/or successional colonisation of duckweed mats.     

The effect of cumulative endurance exercise on leptin and adiponectin and their role as markers to monitor training load

Leptin and adiponectin play an essential role in energy metabolism. Leptin has also been proposed as a marker for monitoring training load. So far, no studies have investigated the variability of these hormones in athletes and how they are regulated during cumulative exercise. This study monitored leptin and adiponectin in 15 endurance athletes twice daily in the days before, during and after a 9-day simulated cycling stage race. Adiponectin significantly increased during the race (p = 0.001) and recovery periods (p = 0.002) when compared to the baseline, while leptin decreased significantly during the race (p < 0.0001) and returned to baseline levels during the recovery period. Intra-individual variability was substantially lower than inter-individual variability for both hormones (leptin 34.1 vs. 53.5%, adiponectin 19% vs. 37.2%). With regards to exercise, this study demonstrated that with sufficient, sustained energy expenditure, leptin concentrations can decrease within the first 24 hours. Under the investigated conditions there also appears to be an optimal leptin concentration which ensures stable energy homeostasis, as there was no significant decrease over the subsequent race days. In healthy endurance athletes the recovery of leptin takes 48-72 hours and may even show a supercompensation-like effect. For adiponectin, significant increases were observed within 5 days of commencing racing, with these elevated values failing to return to baseline levels after 3 days of recovery. Additionally, when using leptin and adiponectin to monitor training loads, establishing individual threshold values improves their sensitivity.     

Capillary electrophoresis for the characterization of quantum dots after non-selective or selective bioconjugation with antibodies for immunoassay

Capillary electrophoresis coupled with laser-induced fluorescence was used for the characterization of quantum dots and their conjugates to biological molecules. The CE-LIF was laboratory-built and capable of injection (hydrodynamic and electrokinetic) from sample volumes as low as 4 μL via the use of a modified micro-fluidic chip platform. Commercially available quantum dots were bioconjugated to proteins and immunoglobulins through the use of established techniques (non-selective and selective). Non-selective techniques involved the use of EDCHCl/sulfo-NHS for the conjugation of BSA and myoglobin to carboxylic acid-functionalized quantum dots. Selective techniques involved 1) the use of heterobifunctional crosslinker, sulfo-SMCC, for the conjugation of partially reduced IgG to amine-functionalized quantum dots, and 2) the conjugation of periodate-oxidized IgGs to hydrazide-functionalized quantum dots. The migration times of these conjugates were determined in comparison to their non-conjugated QD relatives based upon their charge-to-size ratio values. The performance of capillary electrophoresis in characterizing immunoconjugates of quantum dot-labeled IgGs was also evaluated. Together, both QDs and CE-LIF can be applied as a sensitive technique for the detection of biological molecules. This work will contribute to the advancements in applying nanotechnology for molecular diagnosis in medical field.     

Genetic variation and recombination of RdRp and HSP 70h genes of Citrus tristeza virus isolates from orange trees showing symptoms of citrus sudden death disease

Background

Caesarean section before labor or before ruptured membranes ("elective caesarean section", or ECS) has been introduced as an intervention for preventing mother-to-child transmission (MTCT) of hepatitis B virus (HBV). Currently, no evidence that ECS versus vaginal delivery reduces the rate of MTCT of HBV has been generally provided. The aim of this review is to assess, from randomized control trails (RCTs), the efficacy and safety of ECS versus vaginal delivery in preventing mother-to-child HBV transmission.

Results

We searched Cochrane Pregnancy and Childbirth Group's Trials Register (January, 2008), the Cochrane Central Register of Controlled Trials (the Cochrane Library 2008, issue 1), PubMed (1950 to 2008), EMBASE (1974 to 2008), Chinese Biomedical Literature Database (CBM) (1975 to 2008), China National Knowledge Infrastructure (CNKI) (1979 to 2008), VIP database (1989 to 2008), as well as reference lists of relevant studies. Finally, four randomized trails involving 789 people were included. Based on meta-analysis, There was strong evidence that ECS versus vaginal delivery could effectively reduce the rate of MTCT of HBV (ECS: 10.5%; vaginal delivery: 28.0%). The difference between the two groups (ECS versus vaginal delivery) had statistical significance (RR 0.41, 95% CI 0.28 to 0.60, P < 0.000001). No data regarding maternal morbidity or infant morbidity according to mode of delivery were available.

Conclusion

ECS appears to be effective in preventing MTCT of HBV and no postpartum morbidity (PPM) was reported. However, the conclusions of this review must be considered with great caution due to high risk of bias in each included study (graded C).     

Phylogenetic relationships among prokaryotic and eukaryotic catalases

Seventy-four catalase protein sequences, including 29 bacterial, 8 fungal, 7 animal, and 30 plant sequences, were compiled, and 70 were used for phylogenetic reconstruction. The core of the resulting tree revealed unique, separate groups of plant and animal catalases, two groups of fungal catalases, and three groups of bacterial catalases. The only overlap of kingdoms occurred within one branch and involved fungal and bacterial large-subunit enzymes. The other fungal branch was closely linked to the group of animal enzymes. Group I bacterial catalases were more closely related to the plant enzymes and contained such diverse taxa as the Gram-positive Listeria seeligeri, Deinocococcus radiodurans, and gamma-proteobacteria. Group III bacterial sequences were more closely related to fungal and animal sequences and included enzymes from a broad range of bacteria including high- and low-GC Gram positives, proteobacteria, and a bacteroides species. Group II was composed of large-subunit catalases from diverse sources including Gram positives (low-GC Bacilli and high-GC Mycobacteria), proteobacteria, and species of the filamentous fungus Aspergillus. These data can be interpreted in terms of two gene duplication events that produced a minimum of three catalase gene family members that subsequently evolved in response to environmental demands. Horizontal gene transfer may have been responsible for the group II mixture of bacterial and fungal large-subunit catalases.      

The impact of the neisserial DNA uptake sequences on genome evolution and stability

Background  

Efficient natural transformation in Neisseria requires the presence of short DNA uptake sequences (DUSs). Doubts remain whether DUSs propagate by pure selfish molecular drive or are selected for 'safe sex' among conspecifics.     

Unexpected photobleaching of Alexa 488 in a fixed bacterial sample during 2-photon excitation

A sample of fixed bacterial cells was examined by immunofluorescence microscopy using an Alexa 488 conjugated secondary antibody for visualization. Excitation using visible light confirmed the expected photostability of this fluorophore; however, when using 2-photon excitation, Alexa 488 was rapidly and substantially photobleached. The unexpected instability of Alexa 488 under certain conditions may have deleterious consequences if not anticipated and accommodated in experimental protocols.     

Viunalikeviruses are environmentally common agents of horizontal gene transfer in pathogens and biocontrol bacteria

Bacteriophages have been used as natural biocontrol and therapeutic agents, but also as biotechnological tools for bacterial engineering. We showed recently that the transducing bacteriophage ϕMAM1 is a ViI-like phage and a member of the new genus, ‘Viunalikevirus''. Here, we show that four additional ViI-like phages and three new environmentally isolated viunalikeviruses, all infecting plant and human pathogens, are very efficient generalised transducers capable of transducing chromosomal markers at frequencies of up to 10⁻⁴ transductants per plaque-forming unit. We also demonstrate the interstrain transduction of plasmids and chromosomal markers, including genes involved in anabolism, genes for virulence and genes encoding secondary metabolites involved in biocontrol. We propose that all viunalikeviruses are likely to perform efficient horizontal gene transfer. Viunalikeviruses therefore represent useful agents for functional genomics and bacterial engineering, and for chemical and synthetic biology studies, but could be viewed as inappropriate choices for phage therapy.Combined morphological, genomic and phylogenetic analyses have recently led to the proposed creation of a new phage genus, ‘Viunalikevirus'', within the Myoviridae family (Adriaenssens et al., 2012a). The first member of this proposed genus, Salmonella phage ViI, was isolated in the 1930s (Craigie and Yen, 1938) and multiple viunalikeviruses have been sequenced and characterised since 2010 (Pickard et al., 2010; Anany et al., 2011; Hooton et al., 2011; Kutter et al., 2011; Matilla and Salmond, 2012; Park et al., 2012; Adriaenssens et al., 2012a, 2012b; Hsu et al., 2013; Luna et al., 2013; Shahrbabak et al., 2013). Viunalikeviruses are characterised as virulent (lytic) phages showing similar genome size, extensive DNA homology, strong gene synteny and a complex adsorption apparatus, which uses tail spike proteins as host-recognition determinants (Adriaenssens et al., 2012a).We recently isolated the ViI-like phage, ϕMAM1, that infects several environmental and clinical isolates belonging to Serratia and Kluyvera genera (Matilla and Salmond, 2012). During the characterisation of ϕMAM1, we showed that it mediates highly efficient generalised transduction (Matilla and Salmond, submitted for publication). These observations were consistent with a previous report, that the Salmonella phage ViI was also capable of transduction (Cerquetti and Hooke, 1993) and we have confirmed that phage ViI can transduce chromosomal markers and plasmids at frequencies of up to 4.6 × 10⁻⁵ transductants per plaque-forming unit (p.f.u.; Figure 1a; Supplementary Table 1).Open in a separate windowFigure 1Transduction capabilities of viunalikeviruses. (a) Transduction frequencies of LIMEstone1, LIMEstone2, ViI and CBA120 phages. The graph also shows transduction efficiencies of LIMEstone phages within and between Dickeya solani strains. Transduction efficiency was defined as the number of transductants obtained per p.f.u. In all cases, error bars represent the standard deviations (n=3). (b) Skimmed milk agar plates showing protease production in the wild-type (wt) Dickeya solani strains MK10, MK16 and IPO 2222. LIMEstone1- (LS1) and LIMEstone2- (LS2) mediated transduction of the spp::Km marker from the protease negative mutant strain MK10P1 to the wild-type strains MK10, MK16 and IPO 2222 result in a protease-negative phenotype. (c–e) LIMEstone-mediated transduction of the oocN::Km marker from the oocydin A-negative mutant strain MK10oocN to the wild-type strains MK10 (c), MK16 (d) and IPO 2222 (e) results in an oocydin A-negative phenotype and, consequently, in the generation of strains defective in their antimicrobial activity against the plant pathogenic oomycete, Pythium ultimum. The anti-oomycete assays were performed as described previously (Matilla et al., 2012).Most generalised transducers utilise a headful packing strategy where phage terminases recognise specific sequences (pac sites) in the DNA and perform cycles of packing that result in mature phage particles (Fineran et al., 2009a). Indeed, phage terminases with reduced specificity for pac sequences may lead to the evolution of efficient transducing phages (Schmeiger, 1972). Based on the high similarity between the terminases of ϕMAM1, ViI and those of other previously sequenced viunalikeviruses, we hypothesised that all of these ViI-like phages should be capable of transduction in their respective bacterial hosts. To test this hypothesis, we investigated three additional viunalikeviruses, Escherichia coli phage CBA120 (Kutter et al., 2011), and Dickeya phages LIMEstone1 and LIMEstone2 (Adriaenssens et al., 2012b). All the bacteriophages, bacterial strains, plasmids and primers used in this study are listed in the Supplementary Tables 2 and 3. Experimental procedures are presented as Supplementary Material.The LIMEstone phages specifically infect some strains of the emerging plant pathogen, Dickeya solani (Adriaenssens et al., 2012b), and here we showed that they also infect the recently sequenced D. solani strains MK10, MK16 and IPO 2222. As predicted, we confirmed that the LIMEstone phages effected efficient transduction of various auxotrophic markers between Dickeya solani strains (Figure 1a; Supplementary Table 4). To our knowledge, only one Dickeya transducing phage, ϕEC2, has been isolated previously (Resibois et al., 1984). Additional mutant strains were constructed and the generalised nature of the transduction was confirmed by transfer of multiple chromosomal markers, including mutations in the gene cluster encoding biosynthesis of the anti-oomycete haterumalide, oocydin A (Matilla et al., 2012) and in the locus for synthesis and secretion of protease virulence factors. Transduction frequency was higher at an multiplicity of infection (m.o.i.) of 0.1 and 0.01 with efficiencies of up to 10⁻⁴ transductants per p.f.u. (Figure 1a; Supplementary Tables 4 and 5).We also demonstrated transduction of a kanamycin resistance-marked plasmid pECA1039-Km3 between strains MK10, MK16 and IPO 2222 at frequencies of up to 8.6 × 10⁻⁵ (Supplementary Table 4). Plasmid pECA1039 (originally isolated from the phytopathogen, Pectobacterium atrosepticum) encodes a bifunctional type III Toxin-Antitoxin (TA) system, ToxIN, with abortive infection capacity. Although ToxIN aborts infection of various enterobacteria by diverse phages (Fineran et al., 2009b) it did not protect against infection by the tested viunalikeviruses, ϕMAM1, ViI, CBA120, LIMEstone1 or LIMEstone2 (not shown). Furthermore, another type III TA system, TenpIN, from the insect pathogen, Photorhabdus luminescens (Blower et al., 2012), failed to protect against any of the five ViI-like phages (not shown).In addition, we also tested the transduction capacity of the E. coli phage, CBA120, and confirmed transduction of plasmid-borne antibiotic resistances at a frequency of up to 10⁻⁴ transductants per p.f.u. (Figure 1a; Supplementary Table 6).We decided to test our hypothesis that the viunalikeviruses may all be generalised transducers by first isolating new viunalikeviruses from the environment. From treated sewage effluent, we isolated three new bacteriophages infecting Dickeya solani, ϕXF1, ϕXF3 and ϕXF4, as defined initially by their very characteristic ViI-like morphology in electron microscopy (Figures 2a–c). As predicted, all of these new phages were able to transduce chromosomal markers and plasmids at frequencies of up to 3 × 10⁻⁶ transductants per p.f.u. (Figure 2e; Supplementary Table 7). Sequencing of structural and non-structural protein-encoding genes of ϕXF1, ϕXF3 and ϕXF4 showed high nucleotide homology (between 80% and 100%) with the corresponding orthologs in LIMEstone1 (Supplementary Figure 1), indicating that these virgin environmental isolates also clade within the Viunalikevirus genus.Open in a separate windowFigure 2Environmental isolation and characterisation of new viunalikeviruses with generalised transduction functionality. Transmission electron micrographs of phages ϕXF1 (a), ϕXF3 (b), ϕXF4 (c) and ϕXF28 (d) are shown. As an internal control, ϕXF28 was an example of a new lytic phage isolated from the same environment but showing no transduction capabilities. Bars, 50 nm. (e) Transduction frequencies of the new viunalikeviruses ϕXF1, ϕXF3 and ϕXF4. Transduction experiments were performed using 10⁹ cells with ϕXF1, ϕXF3, ϕXF4 at an m.o.i. of 0.01. Transduction efficiency was defined as the number of transductants obtained per p.f.u. Error bars represent the standard deviations (n=3).Although we did not have access to other ViI-like Salmonella phages SFP10 (Park et al., 2012), ϕSH19 (Hooton et al., 2011) and Marshall (Luna et al., 2013), Escherichia phage PhaxI (Shahrbabak et al., 2013), Shigella phage ϕSboM-AG3 (Anany et al., 2011) and Klebsiella phage 0507-KN2-1 (Hsu et al., 2013), our results allow us to predict that all of these phages will mediate generalised transduction. Importantly, these phages would be expected to contribute to the horizontal gene transfer of virulence factors and antimicrobial-resistance determinants in diverse environments.Viunalikeviruses do not seem to be limited to the enterobacteria as bacteriophages showing ViI-like morphology have been isolated in Acinetobacter (Ackermann et al., 1994), Bordetella (Adriaenssens et al., 2012b) and Sinorhizobium (Werquin et al., 1988). Furthermore, another ViI-like morphotype phage (ϕM12 of Sinorhizobium meliloti) has also been shown to be an efficient transducer (Finan et al., 1984). Taken together, these results suggest that, even in the absence of strongly predictive comparative genomic detail, a characteristically discrete ViI-like morphology in electron microscopy may be sufficient to identify new phages as strong candidates for possession of generalised transduction capacity.The emergence and dissemination of antibiotic-resistant pathogens coupled with low discovery rates for new antimicrobials, plus increasing legal constraints on the use of chemical pesticides, have (re)focussed attention on the potential use of bacteriophages for ‘natural biocontrol'' of human, animal and plant pathogens. Several viunalikeviruses have been proposed as candidate therapeutic agents for the control of bacterial infections (Anany et al., 2011; Hooton et al., 2011; Park et al., 2012; Hsu et al., 2013; Shahrbabak et al., 2013) and the LIMEstone phages have been used in successful field trials for biocontrol of D. solani infections (Adriaenssens et al., 2012b). However, their efficient transduction capacities could provide a route for dissemination of virulence factors, such as proteases (Marits et al., 1999). In fact, we have demonstrated the interstrain transduction of plasmids and oocydin A, auxotrophy and protease markers between three different D. solani strains, at high frequencies (Figures 1 and ​and2;2; Supplementary Tables 4 and 7). Also, the irregular distribution of the oocydin A gene cluster within the Dickeya genus and the fact that its genomic context varies between strains raises the possibility of phage-mediated horizontal gene transfer between bacterial strains. These results emphasize strongly that when considering the genomics of phages for ‘phage therapy'' the absence of genes readily defined as playing roles in lysogeny or bacterial virulence may be insufficient to inspire confidence that use of a particular therapeutic phage presents no risk–particularly among the high efficiency-transducing viunalikeviruses.     

In utero protein restriction causes growth delay and alters sperm parameters in adult male rats

Background

Hyperglycemia can impair the male reproductive system in experimental animals and in men during reproductive age. Studies have shown that vitamin C has some good effects on male reproductive system, and therefore vitamin C treatment could attenuate the dysfunctions in this system caused by hyperglycemia. Thus, the objective of this work was to evaluate whether vitamin C treatment could attenuate reproductive dysfunctions in hyperglycemic male rats.

Methods

Adult male rats were divided into 3 groups: a normoglycemic (n = 10) and two hyperglycemic (that received a single dose of streptozotocin - 40 mg/kg BW). The two last groups (n = 10 per group) were divided into: hyperglycemic control (Hy) and hyperglycemic + 150 mg of vitamin C (HyC), by gavage during 30 consecutive days. The normoglycemic and hyperglycemic control groups received the vehicle (water). The first day after the treatment, the rats were anesthetized and killed to evaluate oxidative stress biomarkers (TBARS, SOD, GSHt and GSH-Px) in the erythrocytes, body and reproductive organ weights, sperm parameters, plasma hormone levels (FSH, LH and testosterone), testicular and epididymal histo-morphometry and histopathology.

Results

Compared with the normoglycemic animals, hyperglycemic control rats showed reduced weight of the body and reproductive organ but testis weight was maintained. It was also observed reduction of testosterone and LH levels, seminiferous tubular diameter, sperm motility and sperm counts in the epididymis. In addition, there was an increase in morphological abnormalities on spermatozoa as well as in oxidative stress level. Vitamin C reduced the oxidative stress level, diminished the number of abnormal sperm, and increased testosterone and LH levels and seminiferous tubular diameter but did not show improvement of sperm motility in relation to the hyperglycemic control group. Hyperglycemia caused a rearrangement in the epididymal tissue components (stroma, ephitelium and lumen) as demonstrated by the stereological analysis results. However, this alteration was partially prevented by vitamin C treatment.

Conclusions

We conclude that vitamin C partially attenuated some male reproductive system dysfunctions in hyperglycemic rats.