A solution for private assessment in indirect reciprocity using solitary observation.

Although indirect reciprocity is a fundamental mechanism in the evolution of human cooperation, most studies assume public assessment in which individuals are not permitted to obtain private assessments of others. Existing studies on private assessment have used individual-based simulations because of the analytical difficulty involved. Here, we develop an analytical method using solitary observation to solve private assessment in indirect reciprocity problem without any approximation. In this study, we formulate a model of solitary observation and calculate the replicator dynamics systems of five leading norms of indirect reciprocity. Indirect reciprocity in private assessment provides a different result to that in public assessment. According to the existence proofs of cooperative evolutionarily stable (CES) points in the system, strict norms (stern judging and shunning) have no CES point in private assessment, while they do in public assessment. Image scoring does not change the system regardless of the assessment types because it does not use second-order information. In tolerant norms (simple standing and staying), the CES points move to co-existence of norms and unconditional cooperators. Despite the fact that there is no central coercive assessment system in private assessment, the average cooperation rate at the CES points in private assessment is greater than that in public assessment. This is because private assessment gives unconditional cooperators a role. Our results also show the superiority of the staying norm. Compared with simple standing, staying has three advantages in private assessment: a higher cooperation rate, easiness of invasion into defectors, and robustness to maintain cooperative evolutionarily stable situations. Our results are applicable to general social dilemmas in relation to private information. Under some dilemmas, norms or assessment rules should be carefully chosen to enable cooperation to evolve.

Indirect reciprocity can overcome free-rider problems on costly moral assessment.

Indirect reciprocity is one of the major mechanisms of the evolution of cooperation. Because constant monitoring and accurate evaluation in moral assessments tend to be costly, indirect reciprocity can be exploited by cost evaders. A recent study crucially showed that a cooperative state achieved by indirect reciprocators is easily destabilized by cost evaders in the case with no supportive mechanism. Here, we present a simple and widely applicable solution that considers pre-assessment of cost evaders. In the pre-assessment, those who fail to pay for costly assessment systems are assigned a nasty image that leads to them being rejected by discriminators. We demonstrate that considering the pre-assessment can crucially stabilize reciprocal cooperation for a broad range of indirect reciprocity models. In particular for the most leading social norms, we analyse the conditions under which a prosocial state becomes locally stable.

Prevalence and first genotyping of Giardia duodenalis in beef calves in Vietnam.

Little information is available on the epidemiology of Giardia duodenalis in beef cattle from Vietnam. This study was performed to determine the prevalence and genotypes/assemblages of G. duodenalis in native beef calves younger than 6 months in the region. A total of 412 calf fecal samples, randomly selected from 99 small-scale farms located in DacLac and KhanhHoa provinces, central Vietnam, were screened for the presence of G. duodenalis cysts using the zinc-sulfate flotation method followed by iodine staining. The overall prevalence on the sample and herd levels were 13.8% (57/412) and 42.4% (42/99), respectively. Molecular analysis in the ß-giardin and triosephosphate isomerase genes demonstrated the presence of only G. duodenalis assemblage E in the animals. Since assemblage E has been rarely reported in humans, the zoonotic risk in beef calves in the region appears to be minimal.

Role of the Cytosolic Heat Shock Protein 70 Ssa5 in the Ciliate Protozoan Tetrahymena thermophila.

Heat shock protein 70 (Hsp70) is a member of a family of conserved chaperone proteins whose function is well investigated in many model organisms. Here we focus on an Hsp70 called Ssa5 in the ciliate protozoan Tetrahymena thermophila, and reveal that its translation is heat inducible as for general Hsps. Moreover, the protein is abundantly expressed in the cytoplasm during sexual reproduction (conjugation) as well as in response to heat-stress. Knocking out of SSA5 (ΔSSA5) does not affect the survival of the cell under heat-stress, likely due to other Hsp70 paralogs compensating for the defect. During conjugation, ΔSSA5 leads to a fertilization defect in which the two pronuclei are in close proximity but never fuse. The unfertilized pronuclei differentiate, resulting in a heterokaryon with developed haploid germline and somatic nuclei. In addition, degeneration of the parental somatic nucleus is not affected. These results suggest a specific involvement of Ssa5 in pronuclear fusion and fertilization.

Seawater inundation from the 2011 Tohoku tsunami continues to strongly affect soil bacterial communities 1 year later.

The effects of inundation caused by the 2011 Tohoku tsunami on soil bacterial communities in agricultural fields were evaluated. Bacterial communities were compared across three different types of soil, unflooded field (UF) soil, soil flooded for 2 weeks (short term (ST)), and soil flooded for 2 months (long term (LT)), using polymerase chain reaction-pyrosequencing of 16S rRNA genes. Acidobacteria were dominant in UF, with a relative abundance of approximately 35 %, and Proteobacteria dominated flooded soils (30-67 %). Hierarchical cluster analysis indicated that the community structure of soil bacteria in flooded soils (ST and LT) clearly differed from that in UF. Differences between LT and ST fields were rarely observed in terms of chemical properties and microbial community structure at the phylum level. However, sulfur-oxidizing bacteria (SOB) and nitrite-oxidizing bacteria (NOB) in LT tended to occur at high and low abundances, respectively. Halothiobacillus, a halotolerant SOB, was detected in all LT fields. Unexpectedly, a zeta-Proteobacteria, which had previously only been detected in marine environments, was detected in LT fields only. Our results demonstrate that the effects of the 2011 Tohoku tsunami on soil bacterial communities in agricultural fields may have lasted at least 1 year. Furthermore, SOB, NOB, and zeta-Proteobacteria may serve as indicators of the effects of seawater inundation on microorganisms.

Pathological features of Cryptosporidium andersoni-induced lesions in SCID mice.

To assess the infectivity and the istopathological features of Cryptosporidium andersoni (C. andersoni) in laboratory animals, SCID mice were orally inoculated with oocysts of C. andersoni. Starting one week after inoculation, the SCID mice began shedding oocysts, and this continued for ten weeks. Histopathologically, myriads of C. andersoni were observed on the apical surface of the epithelium in the gastric pit of the glandular stomach. There were few lesions in the gastric epithelium except C. andersoni adhesion. In the lamina propria of the affected mucosa, minimum infiltration of inflammatory cells was observed. Immunohistochemically, C. andersoni demonstrated a positive reaction to a number of primary antibodies of Cryptosporidium parvum. In the experiment described here, few increases were seen in apoptotic epithelial cells in the affected mucosas of the SCID mice, and the nuclear augmentation was not enhanced. It was hypothesized that the absence of apoptosis and cell division were due to a lack of inflammatory cell reaction in the lamina propria.

Prevalence and molecular characterization of Cryptosporidium in ostriches (Struthio camelus) on a farm in central Vietnam.

This study was performed to determine the prevalence and molecular characterization of Cryptosporidium in ostriches on a farm in Khanh Hoa province, central Vietnam. A total of 464 ostrich fecal samples were examined Cryptosporidium oocysts using the modified Ziehl-Neelsen staining method, and 110 (overall prevalence 23.7%) were identified as positive by microscopy. Prevalence of Cryptosporidium in animals of <45 days, 45-60 days, 61-90 days, 91 days-12 months and >12 months was 23.5% (16/68), 33.3% (22/66), 35.2% (68/193), 0 and 5.8% (4/69), respectively (p<0.05). The majority of positive samples scored as the 3+ level of intensity of infection were from 61 to 90 days ostriches. Molecular analysis in the 18S ribosomal RNA, 70 kDa heat shock protein and actin genes demonstrated the presence of only Cryptosporidium avian genotype II in ostriches in central Vietnam.

Molecular characterization of Cryptosporidium in pigs in central Vietnam.

Little information is available on the epidemiology of Cryptosporidium in pigs in central Vietnam. The aims of this study were to investigate the prevalence and to characterize the genotype distribution of Cryptosporidium isolates in pigs in this region. A total of 193 pig fecal samples were screened for the presence of Cryptosporidium oocysts using the modified Ziehl-Neelsen staining method, and 28 (overall prevalence 14.5 %) were identified as positive by microscopic observation. Positive samples were further analyzed by polymerase chain reaction amplification and sequencing. Genetic identification based on the 18S ribosomal RNA and 70 kDa heat shock protein genes revealed that pigs in Vietnam are infected with two species/genotypes (Cryptosporidium suis and Cryptosporidium pig genotype II). This study is the first molecular characterization of Cryptosporidium in pigs in Vietnam. The presence of these host-adapted species/genotypes suggests that pigs may not pose a significant public health risk in this area. More extensive studies are necessary to ascertain the zoonotic potential of Cryptosporidium in porcine hosts in Vietnam.

Relationship Between Rumen Microbial Composition and Fibrolytic Isozyme Activity During the Biodegradation of Rice Straw Powder Using Rumen Fluid.

Rumen fibrolytic microorganisms have been used to increase the rate of lignocellulosic biomass biodegradation; however, the microbial and isozymatic characteristics of biodegradation remain unclear. Therefore, the present study investigated the relationship between rumen microorganisms and fibrolytic isozymes associated with lignocellulosic biomass hydrolysis. Rice straw, a widely available agricultural byproduct, was ground and used as a substrate. The biodegradation of rice straw powder was performed anaerobically in rumen fluid for 48 h. The results obtained revealed that 31.6 and 23.3% of cellulose and hemicellulose, respectively, were degraded. The total concentration of volatile fatty acids showed a 1.8-fold increase (from 85.4 to 151.6| |mM) in 48 h, and 1,230.1| |mL L-1 of CO2 and 523.5| |mL L-1 of CH4 were produced. The major isozymes identified by zymograms during the first 12| |h were 51- and 140-kDa carboxymethyl cellulases (CMCases) and 23- and 57-kDa xylanases. The band densities of 37-, 53-, and 58-kDa CMCases and 38-, 44-, and 130-kDa xylanases increased from 24 to 36 h. A microbial ana-lysis indicated that the relative abundances of Prevotella, Fibrobacter, and Bacteroidales RF16 bacteria, Neocallimastix and Cyllamyces fungi, and Dasytricha and Polyplastron protozoa were related to fibrolytic isozyme activity. The present results provide novel insights into the relationships between fibrolytic isozymes and rumen microorganisms during lignocellulose biodegradation.

Growth of ammonia-oxidizing archaea and bacteria in cattle manure compost under various temperatures and ammonia concentrations.

A recent study showed that ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) coexist in the process of cattle manure composting. To investigate their physiological characteristics, liquid cultures seeded with fermenting cattle manure compost were incubated at various temperatures (37°C, 46°C, or 60°C) and ammonium concentrations (0.5, 1, 4, or 10 mM NH (4) (+) -N). The growth rates of the AOB and AOA were monitored using real-time polymerase chain reaction analysis targeting the bacterial and archaeal ammonia monooxygenase subunit A genes. AOB grew at 37°C and 4 or 10 mM NH (4) (+) -N, whereas AOA grew at 46°C and 10 mM NH (4) (+) -N. Incubation with allylthiourea indicated that the AOB and AOA grew by oxidizing ammonia. Denaturing gradient gel electrophoresis and subsequent sequencing analyses revealed that a bacterium related to Nitrosomonas halophila and an archaeon related to Candidatus Nitrososphaera gargensis were the predominant AOB and AOA, respectively, in the seed compost and in cultures after incubation. This is the first report to demonstrate that the predominant AOA in cattle manure compost can grow and can probably oxidize ammonia under moderately thermophilic conditions.

Molecular characterization of Cryptosporidium in native beef calves in central Vietnam.

The aims of this study were to investigate the prevalence of Cryptosporidium and to characterize the genotype distribution of Cryptosporidium isolates in native beef calves 2-6 months old in Dac Lac province, central Vietnam. The presence of Cryptosporidium oocysts was determined using the modified Ziehl-Neelsen staining method. The overall prevalence on the sample and herd levels were 18.9% (44/232) and 50% (20/40), respectively. Genotyping based on PCR and sequence analysis of the 18 S rRNA gene revealed occurrence of the two nonzoonotic species Cryptosporidium ryanae and Cryptosporidium bovis, with the former as a dominant species in the animals. The absence of the zoonotic species Cryptosporidium parvum in calves examined suggests that the native beef calves 2-6 months old in the study area are unlikely to contribute to human cryptosporidiosis transmission.

Middle-thermophilic sulfur-oxidizing bacteria Thiomonas sp. RAN5 strain for hydrogen sulfide removal.

Hydrogen sulfide (H2S) is one of the most toxic and offensively odorous gases and is generated in anaerobic bioreactors. A middle-thermophilic sulfur-oxidizing bacterium (SOB), Thiomonas sp. strain RAN5, was isolated and applied for H2S removal from both artificial and anaerobically digested gas. When a bioreactor containing medium inoculated with RAN5 was aerated continuously with artificial gas (containing 100 ppm H2S) at 45 degrees C for 156 hr, the H2S concentration in the vented gas was reduced by 99%. This was not affected by the presence of other microbes in the bioreactor The H2S removal efficiency of the RAN5 bioreactor for anaerobically digested gas was greater than 99% at influent H2S concentrations ranging from 2 to 1800 ppm; the efficiency decreased to 90% at influent H2S concentrations greater than 2000 ppm. Thiomonas sp. strain RAN5 cannot survive at room temperature, and thus its leakage from a wastewater treatment plant would not damage sewage systems. These data suggest that Thiomonas sp. strain RAN5 may be a useful microorganism for H2S removal.

Prevalence of Fasciola in cattle and of its intermediate host Lymnaea snails in central Vietnam.

The aims of this study were to investigate the prevalence of natural Fasciola infections in both the definitive hosts (cattle) and the intermediate hosts (Lymnaea snails) in central Vietnam. A total of 1,075 fecal samples, randomly collected from cattle in Binh Dinh, Khanh Hoa, and Phu Yen provinces, were examined for Fasciola eggs by a sedimentation method. The overall prevalence of Fasciola was 45.3 %. A subset of the animals (235) was also screened for antibodies against Fasciola by an enzyme-linked immunosorbent assay. Overall, 46.3 % of these animals were shedding Fasciola eggs while 87.2 % were Fasciola seropositive. A lower prevalence of Fasciola was observed in calves ≤ 2 years of age (37.6 %) compared to that in cattle >2 years of age (53.7 %) (p < 0.05). The prevalence in the rainy season (50.8 %) was significantly different to that in the dry season (38.1 %) (p < 0.05). Of the 3.269 Lymnaea viridis and 1.128 Lymnaea swinhoei examined, 31 (0.95 %) and seven (0.62 %), respectively, were found to be infected with Fasciola. This appears to be the first epidemiological survey of the prevalence of Fasciola in cattle and snails in these three provinces in central Vietnam.

Shifts in xylanases and the microbial community associated with xylan biodegradation during treatment with rumen fluid.

Treatment with rumen fluid improves methane production from non-degradable lignocellulosic biomass during subsequent methane fermentation; however, the kinetics of xylanases during treatment with rumen fluid remain unclear. This study aimed to identify key xylanases contributing to xylan degradation and their individual activities during xylan treatment with bovine rumen microorganisms. Xylan was treated with bovine rumen fluid at 37°C for 48 h under anaerobic conditions. Total solids were degraded into volatile fatty acids and gases during the first 24 h. Zymography showed that xylanases of 24, 34, 85, 180, and 200 kDa were highly active during the first 24 h. Therefore, these xylanases are considered to be crucial for xylan degradation during treatment with rumen fluid. Metagenomic analysis revealed that the rumen microbial community's structure and metabolic function temporally shifted during xylan biodegradation. Although statistical analyses did not reveal significantly positive correlations between xylanase activities and known xylanolytic bacterial genera, they positively correlated with protozoal (e.g., Entodinium, Diploplastron, and Eudiplodinium) and fungal (e.g., Neocallimastix, Orpinomyces, and Olpidium) genera and unclassified bacteria. Our findings suggest that rumen protozoa, fungi, and unclassified bacteria are associated with key xylanase activities, accelerating xylan biodegradation into volatile fatty acids and gases, during treatment of lignocellulosic biomass with rumen fluid.

Accurate analysis of prevalence of coccidiosis in individually identified wild cranes in inhabiting and migrating populations in Japan.

Eimeria gruis and E. reichenowi cause coccidiosis, a major parasitic disease of cranes. By non-invasive molecular approaches, we investigated the prevalence and genetic characterization of pathogens in two Japanese crane habitats; one is Hokkaido inhabited by the endangered red-crowned crane, and the other is Izumi in Kyushu where populations that consist mainly of vulnerable hooded and white-naped cranes migrate in winter. The non-invasively collected faecal samples from each wintering population were first subjected to host genomic DNA-targeted analyses to determine the sample origin and avoid sample redundancy. Extremely high prevalence was observed in the Izumi populations (> 90%) compared with the Hokkaido population (18-30%) by examining 470 specimens by microscopy and PCR-based capillary electrophoresis (PCR-CE), using genetic markers in the second internal transcribed spacer (ITS2). Correspondence analysis of PCR-CE data revealed differences in community composition of coccidia between hooded and white-naped cranes. 18S rRNA and ITS2 sequences were determined from single oocysts excreted by red-crowned and hooded cranes. Phylogenetic analysis of 18S rRNA suggested that E. reichenowi was polyphyletic while E. gruis was monophyletic. Together with PCR-CE data, these results indicate different host specificity among the E. reichenowi type. Our data suggest that E. reichenowi comprises multiple species.

Archaeal community dynamics and detection of ammonia-oxidizing archaea during composting of cattle manure using culture-independent DNA analysis.

The composting process is carried out under aerobic conditions involving bacteria, archaea, and fungi. Little is known about the diversity of archaeal community in compost, although they may play an important role in methane production and ammonia oxidation. In the present study, archaeal community dynamics during cattle manure composting were analyzed using a clone library of the archaeal 16S rRNA gene. The results indicated that methane-producing archaea (methanogen) and ammonia-oxidizing archaea (AOA) may be the dominant microbes throughout the composting. The community consisted primarily of Methanocorpusculum-like and Methanosarcina-like sequences until day 2, while the number of Candidatus Nitrososphaera-like sequences increased from day 6 to day 30. Methanosarcina thermophila-like sequences were dominant from day 2, suggesting that M. thermophila-like species can adapt to increasing temperature or nutrient loss. A denaturant gradient gel electrophoresis analysis of the archaeal amoA genes revealed that the dominant amoA gene sequence with 99% homology to that of Candidatus Nitrososphaera gargensis was identical to those obtained from a different composting facility. These data suggested that AOA may play a role in ammonia oxidation in several composting practices. Our results provide fundamental information regarding archaeal community dynamics that will help in understanding the collective microbial community in compost.

Detection of Escherichia coli in a cattle manure composting process by selective cultivation and colony polymerase chain reaction.

Livestock manure is suitable for use as a composting material. However, various intestinal microbes, such as Escherichia coli, are significant components of such manures. Thus, it is desirable that the level of intestinal microbes, and particularly opportunistic pathogens, in compost is inspected and counted regularly. The sensitivity and specificity of detection of E. coli in compost have been improved by selective cultivation followed by colony polymerase chain reaction (PCR) using the ECO primer. Indeed, the sensitivity of this method is higher than that of DNA extraction from compost and PCR. In this study, changes in numbers of E. coli present in a field-scale composting process over time was assessed using selective cultivation and colony PCR. Numbers of ECO-positive colonies after 24 h decreased, with a concomitant rise in compost temperature. ECO-positive colonies were not detected from 33 to 48 h. However, ECO-positive colony numbers increased beginning on day 4 and continuing until day 42. Thus, it seems likely that the high temperatures reached during the composting process did not affect E. coli numbers in the final compost. Additionally, selective cultivation followed by colony PCR using specific primers is an appropriate method of determining levels of cultivable pathogens in composted materials.

Characteristics of various fibrolytic isozyme activities in the rumen microbial communities of Japanese Black and Holstein Friesian cattle under different conditions.

Rumen microorganisms produce various fibrolytic enzymes and degrade lignocellulosic materials into nutrient sources for ruminants; therefore, the characterization of fibrolytic enzymes contributing to the polysaccharide degradation in the rumen microbiota is important for efficient animal production. This study characterized the fibrolytic isozyme activities of a rumen microbiota from four groups of housed cattle (1, breeding Japanese Black; 2, feedlot Japanese Black; 3, lactating Holstein Friesian; 4, dry Holstein Friesian). Rumen fluids in all cattle groups showed similar concentrations of total volatile fatty acids and reducing sugars, whereas acetic acid contents and pH were different among them. Predominant genera were commonly detected in all cattle, although the bacterial compositions were different among cattle groups. Zymograms of whole proteins in rumen fluids showed endoglucanase activities at 55 and 57 kDa and xylanase activity at 44 kDa in all cattle. Meanwhile, several fibrolytic isozyme activities differed among cattle groups and individuals. Treponema, Succinivibrio, Anaeroplasma, Succiniclasticum, Ruminococcus, and Butyrivibrio showed positive correlations with fibrolytic isozyme activities. Further, endoglucanase activity at 68 kDa was positively correlated with pH. This study suggests the characteristics of fibrolytic isozyme activities and their correlations with the rumen microbiota.

Diversity and abundance of ammonia-oxidizing bacteria and ammonia-oxidizing archaea during cattle manure composting.

Ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) play important roles in nitrification in various environments. They may also be key communities for ammonia oxidation in composting systems, although few studies have discussed their presence. We investigated the relative diversity and abundance of AOB and AOA using cloning procedures, denaturing gradient gel electrophoresis analysis, and real-time PCR during several stages in the process of cattle manure composting. Our results revealed that the AOB community structure changed during the process. At the high-temperature stage (>60°C), a member of the Nitrosomonas europaea/eutropha cluster dominated while the uncultured Nitrosomonas spp. cluster appeared after the temperature decreased. Additionally, our analysis indicated that AOA sequences, which were classified into a soil/sediment cluster, were present after the temperature decreased during the composting process. At these stages, the number of the archaeal amoA gene copies (3.2 or 3.9 × 10(7) copies per gram freeze-dried compost) was significantly higher than that of bacterial amoA gene copies (2.2-7.2 × 10(6) copies per gram freeze-dried compost). Our results suggest that both AOB and AOA are actively involved in nitrification of composting systems.

Multilocus genetic analysis of Cryptosporidium parvum from Egypt.

Cryptosporidium parvum is a ubiquitous zonootic parasite causing enteritis in man and animals. Cryptosporidium infection was confirmed microscopically in neonatal calves (less than 6 weeks of age) at Kafr El Sheikh Province, Egypt. Multilocus analysis using a wide array of genetic markers was carried out to assess genetic diversity of C. parvum isolates. PCR amplification and partial sequence analysis of 70 kDa heat shock protein, dihydrofolate reductase, alpha-tubulin, elongation factor 1 alpha as well as thrombospondin-related anonymous protein of Cryptosporidium-1, and thrombospondin-related anonymous protein of Cryptosporidium-2 gene markers were achieved. Data indicated that the analyzed isolates belong to C. parvum genotype II with obvious sequence heterogeneity compared with counterparts deposited in Genebank.