A socio-economic study of Fasciola infections in cattle and sheep at the Etoudi slaughterhouse, Yaoundé, Cameroon.

A cross-sectional socio-economic study of fascioliasis in livestock was carried out at the Livestock Development Cooperation (SODEPA) slaughterhouse in Etoudi, Yaoundé during the periods covering the beginning and late dry season in the northern and eastern regions of Cameroon. A total of 768 cattle and 267 sheep were inspected for the presence of Fasciola species. The overall infection rate in cattle and sheep was 18% (n = 767) and 27% (n = 267), respectively. For the animals that were Fasciola positive, a total of 267.86 kg of liver in cattle was condemned, resulting in a loss of US$1124, while a total of 57 kg of liver was condemned from the sheep population, amounting to a loss of US$114. A total of US$76,097 was determined as losses incurred from condemnation of liver for both cattle and sheep per annum based on the total number slaughtered each year. The findings indicate that fascioliasis is present in cattle and sheep slaughtered in Cameroon and that it causes great economic losses due to condemnation of liver. The local climatic conditions, husbandry systems and the presence of snails (intermediate hosts) are probably the main factors influencing the incidence of the disease and may account for the epidemiological significance found in this study.

Molecular identification of bloodmeal sources and trypanosomes in Glossina spp., Tabanus spp. and Stomoxys spp. trapped on cattle farm settlements in southwest Nigeria.

The interactions of host, vector and parasite in bovine trypanosomiasis transmission cycles in southwest Nigeria are not yet well understood. Trypanosoma (Trypanosomatida: Trypanosomatidae) species infection prevalences and bloodmeal sources were determined in transmitting vectors of the genera Glossina (Diptera: Glossinidae), Tabanus (Diptera: Tabanidae) and Stomoxys (Diptera: Muscidae) collected using Nzi traps in cattle settlements in southwest Nigeria. Sequenced cytochrome B mitochondrial DNA segments obtained from vector digestive tracts identified bloodmeal sources from eight host species, namely human, cattle, hippopotamus, giraffe, gazelle, spotted hyena, long-tailed rat and one unidentified species. Overall, 71.1% [95% confidence interval (CI) 63.0-78.1], 33.3% (95% CI 21.9-47.0) and 22.2% (95% CI 16.2-29.9), respectively, of Glossina, Tabanus and Stomoxys flies were positive for trypanosomes. The observed trypanosome species were Trypanosoma vivax, Trypanosoma congolense, Trypanosoma brucei, Trypanosoma evansi, Trypanosoma simiae and Trypanosoma godfreyi. Trypanosome DNA was more prevalent in tsetse (34.8% Tr. vivax, 51.1% Tr. b. brucei, 5.2% Tr. congolense, 4.4% Tr. simiae and 24.4% mixed infections) than in other flies and the main determinants in all flies were seasonal factors and host availability. To the best of the present group's knowledge, this is the first report of Trypanosoma species in Tabanus and Stomoxys flies in Nigeria. It indicates that vector control programmes should always consider biting flies along with tsetse flies in the control of human and animal trypanosomiasis.

People, Patches, and Parasites: The Case of Trypanosomiasis in Zimbabwe.

Understanding the socio-ecology of disease requires careful attention to the role of patches within disease landscapes. Such patches, and the interfaces between different socio-epidemiological systems, we argue, have important implications for disease control. We conducted an interdisciplinary study over three years to investigate the spatial dynamics of human and animal trypanosomiasis in the Zambezi valley, Zimbabwe. We used a habitat niche model to identify changes in suitable habitat for tsetse fly vectors over time, and this is related to local villagers' understandings of where flies are found. Fly trapping and blood DNA analysis of livestock highlighted the patchy distribution of both flies and trypanosome parasites. Through livelihoods analysis we explored who makes use of what areas of the landscape and when, identifying the social groups most at risk. We conclude with a discussion of the practical implications, including the need for an integrated 'One Health' approach involving targeted approaches to both vector control and surveillance.

Expression and Characterization of HA1 Protein of Highly Pathogenic H5N1 Avian Influenza Virus for Use in a Serodiagnostic Assay.

The hemagglutinin ectodomain (HA1 subunit) from highly pathogenic avian influenza (HPAI) isolate (A/chicken/Vietnam/14/2005) was cloned and expressed using a baculovirus expression vector. Biosynthesis, glycosylation and secretion of the HA1 proteins, with natural or a melittin signal peptide at the N-terminus and a six-histidine (6xHis) tag at the C-terminus, were examined in insect cells. A 40-kDa unglycosylated precursor and a fully processed, mature form of the HA1 protein migrated around 52 kDa were detected by SDS-PAGE and confirmed by Western blot using H5N1-specific antibody. Treatment of tunicamycin and peptide-N-glycosidase F (PNGase F) further revealed that the recombinant HA1 proteins produced in insect cells were indeed glycosylated with N-linked oligosaccharide side chains. Time-course experiments showed that substitution of the HA natural sequence with the signal sequence from honeybee melittin promoted a high level of expression and efficient secretion of the HA1. A high yield, 37 µg/ml, of HA1 protein was obtained from recombinant baculovirus-infected cell culture supernatant. In addition, the cell surface expression of rHA1 was detected by indirect immunofluorescent staining and showed biological activity on hemadsorption assays. Recombinant HA1 protein-based ELISA was evaluated and appeared to be sensitive and specific for the rapid detection of H5 subtype-specific antibodies in serum samples. No cross-reactivity to antibodies from 15 other influenza A subtypes was detected. Taken together, the newly developed recombinant HA1-based ELISA could offer an alternative to other diagnostic approaches for the specific detection of H5 avian influenza virus infection.

Socially flexible female choice and premating isolation in field crickets (Teleogryllus spp.).

Social influences on mate choice are predicted to influence evolutionary divergence of closely related taxa, because of the key role mate choice plays in reproductive isolation. However, it is unclear whether females choosing between heterospecific and conspecific male signals use previously experienced social information in the same manner or to the same extent that they do when discriminating among conspecific mates only. We tested this using two field cricket sister species (Teleogryllus oceanicus and Teleogryllus commodus), in which considerable information is known about the role of male calling song in premating isolation, in addition to the influence of acoustic experience on the development of reproductive traits. We manipulated the acoustic experience of replicate populations of both species and found, unexpectedly, that experience of male calling song during rearing did not change how accurate females were in choosing a conspecific over a heterospecific male song during playback trials. However, females with acoustic experience were considerably less responsive to male song compared with naïve females. Our results suggest that variation in the acoustic environment affects mate choice in both species, but that it may have a limited impact on premating isolation. The fact that social flexibility during interspecific mate discrimination does not appear to operate identically to that which occurs during conspecific mate discrimination highlights the importance of considering the context in which animals exercise socially flexible mating behaviours. We suggest an explanation for why social flexibility might be context dependent and discuss the consequences of such flexibility for the evolution of reproductive isolation.

Baculovirus expression and antigenic characterization of classical swine fever virus E2 proteins.

Genes encoding a major structural glycoprotein, E2, of classical swine fever viruses (CSFV) Brescia (subgroup 1.2), Paderborn (subgroup 2.1) and Kanagawa (subgroup 3.4) were constructed by removing the transmembrane domain and adding a C-terminal 6 histidine (His) tag. All the E2 constructs were efficiently expressed in a baculovirus system as 53-kDa glycosylated proteins that were identified in Western blots by their reaction with anti-His and CSFV-specific antibodies. These proteins were used as ELISA antigens to confirm the existence of an antigenic relationship between the viruses using group-specific polyclonal antisera. Antigenic differences were identified by Western blot and ELISA reactivity of the E2 proteins with a panel of monoclonal antibodies. Specifically, one monoclonal antibody (WH303) reacted with all three proteins, two monoclonal antibodies (M1660 and M1665) reacted with only the Brescia E2 protein, and three monoclonal antibodies (M1654, M1664 and M1669) reacted equally well with only Brescia and Kanagawa E2 proteins. Therefore, antibody reactivity profiles, established using recombinant E2 proteins, could be used to quickly identify novel CSFV strains as illustrated in this report with only a limited number of monoclonal antibodies. These proteins could also have added utility in the production of monoclonal antibodies and as critical reagents in diagnostic assays.

Nutritional management of PKU with glycomacropeptide from cheese whey.

Individuals with phenylketonuria (PKU) must follow a lifelong low-phenylalanine (Phe) diet to prevent neurological impairment. Compliance with the low-Phe diet is often poor owing to restriction in natural foods and the requirement for consumption of a Phe-free amino acid formula or medical food. Glycomacropeptide (GMP), a natural protein produced during cheese-making, is uniquely suited to a low-Phe diet because when isolated from cheese whey it contains minimal Phe (2.5-5 mg Phe/g protein). This paper reviews progress in evaluating the safety, acceptability and efficacy of GMP in the nutritional management of PKU. A variety of foods and beverages can be made with GMP to improve the taste, variety and convenience of the PKU diet. Sensory studies in individuals with PKU demonstrate that GMP foods are acceptable alternatives to amino acid medical foods. Studies in the PKU mouse model demonstrate that GMP supplemented with limiting indispensable amino acids provides a nutritionally adequate source of protein and improves the metabolic phenotype by reducing concentrations of Phe in plasma and brain. A case report in an adult with classical PKU who followed the GMP diet for 10 weeks at home indicates safety, acceptability of GMP food products, a 13-14% reduction in blood Phe levels (p<0.05) and improved distribution of dietary protein throughout the day compared with the amino acid diet. In summary, food products made with GMP that is supplemented with limiting indispensable amino acids provide a palatable alternative source of protein that may improve dietary compliance and metabolic control of PKU.

Role of visual and olfactory cues from agricultural hedgerows in the orientation behavior of multicolored Asian lady beetle (Coleoptera: Coccinellidae).

Harmonia axyridis Pallas is an introduced lady beetle common in eastern North American agroecosystems. Two-choice behavioral bioassays were performed to determine whether visual and olfactory stimuli from prey and host habitats could elicit taxis in wild-collected H. axyridis adults and whether beetles exhibit a preference among stimuli. Soybean aphid (Aphis glycines Matsumura) spends much of the year in agricultural hedgerows residing on buckthorn (Rhamnus cathartica L), and H. axyridis is frequently observed feeding on aphids in this habitat. Olfactory bioassays were performed in a Y-tube olfactometer and tested the response of beetles to the odor of buckthorn leaves, apple leaves (Malus domestica Borkh.), and buckthorn leaves both naturally and artificially infested with A. glycines. No differences were observed between the numbers of beetles moving toward the odor of buckthorn artificially infested with A. glycines and uninfested buckthorn, but more beetles preferred naturally infested buckthorn over uninfested buckthorn. Visual bioassays were performed in an acrylic tube arena,and tested beetle response to silhouettes and to apple and buckhorn leaves. Beetles were significantly more likely to choose silhouettes over blank space in visual trials. Significantly more beetles moved toward buckthorn leaves than blank space, but beetles did not discern between apple and buckthorn until olfactory cues were also included. This study lays the foundation for future work examining the response of H. axyridis to visual and olfactory cues in Ontario agroecosystems, which could help enhance effectiveness of H. axyridis as a biological control and mitigate its impacts as a pest species.

Antioxidants promote establishment of trypanosome infections in tsetse.

Efficient, cyclical transmission of trypanosomes through tsetse flies is central to maintenance of human sleeping sickness and nagana across sub-Saharan Africa. Infection rates in tsetse are normally very low as most parasites ingested with the fly bloodmeal die in the fly gut, displaying the characteristics of apoptotic cells. Here we show that a range of antioxidants (glutathione, cysteine, N-acetyl-cysteine, ascorbic acid and uric acid), when added to the insect bloodmeal, can dramatically inhibit cell death of Trypanosoma brucei brucei in tsetse. Both L- and D-cysteine invoked similar effects suggesting that inhibition of trypanosome death is not dependent on protein synthesis. The present work suggests that antioxidants reduce the midgut environment protecting trypanosomes from cell death induced by reactive oxygen species.

Programmed cell death in African trypanosomes.

Until recently it had generally been assumed that apoptosis and other forms of programmed cell death evolved during evolution of the metazoans to regulate growth and development in these multicellular organisms. However, recent research is adding strength to the original phenotypic observations described almost a decade ago which indicated that some parasitic protozoa may have evolved a cell death pathway analogous to the process described as apoptosis in metazoa. Here we explore the implications of a programmed cell death pathway in the African tsetse-transmitted trypanosomes.

Minority populations in the child welfare system: the visibility hypothesis reexamined.

This study tests the "visibility hypothesis," which suggests that there is a higher probability for minority children to be placed in foster care when living in geographic locations where their proportions in the population are relatively low, compared to areas where their proportions are high. Results support the hypothesis for African-American youth only. Explanations of the findings, and their implications for research and practice, are discussed.

Type of maltreatment as a predictor of mental health service use for children in foster care.

This study examined predictors of utilization of mental health services among children and adolescents in foster care. Of particular interest was whether the type of maltreatment was related to utilization of services. Subjects were 662 children ages 2-17 who were in foster care for at least 5 months. Service utilization, demographic, and behavioral data were collected through interviews with the subjects' caretakers. Type of maltreatment data were collected from Child Welfare case records. Fifty-six percent of the subjects had received mental health services. Children removed from their homes due to sexual and/or physical abuse were more likely to receive services than were those who were removed due to neglect and caretaker absence. Clinically significant behavior problems were associated with greater likelihood of receiving services, except for the sexually abused group who were very likely to receive services regardless of their behavior problem score. Sexually abused youth also received a higher number of outpatient visits than did neglected youth. In sum, youth who have experienced "active" types of maltreatment are more likely to receive mental health services than are those with "passive" types of maltreatment, even when the effect of severity of mental health problems is controlled.

Chemical egg defense in a green lacewing (Ceraeochrysa smithi)

The green lacewing Ceraeochrysa smithi (Neuroptera, Chrysopidae), like other members of its family, lays its eggs on stalks, but it is unusual in that it coats these stalks with droplets of an oily fluid. The liquid consists of a mixture of fatty acids, an ester, and a series of straight-chain aldehydes. Relative to the eggs of a congeneric chrysopid that lacks stalk fluid, the eggs of C. smithi proved well protected against ants. Components of the fluid, in an assay with a cockroach, proved potently irritant. Following emergence from the egg, C. smithi larvae imbibe the stalk fluid, thereby possibly deriving nutritive benefit, defensive advantage, or both.

Evaluation of a commercial polymerase chain reaction assay for Chlamydia trachomatis and suggestions for improving sensitivity.

A commercial polymerase chain reaction (PCR) assay (Amplicor, Roche) for Chlamydia trachomatis was compared with a direct fluorescent antibody (DFA) test using urethral and cervical samples, many of which on the basis of prior testing by DFA contained small rather than large numbers of elementary bodies. Urine samples were collected from patients in a sequential unselected manner. Of 244 clinical specimens (138 male urethral and cervical; 106 male and female urine), 66 were positive by both DFA and PCR and 141 were negative by both tests. Nine samples were DFA negative and PCR positive, and 28 samples were DFA positive and PCR negative. However, 24 (86%) of the latter samples contained fewer than ten elementary bodies. When serial dilutions of laboratory stock strains (serovars E and H) were tested, the DFA test detected Chlamydia trachomatis at a dilution tenfold greater than the PCR. Furthermore, of five DFA-positive clinical samples, three that were PCR negative when tested according to the manufacturer's instructions were positive when they were diluted less. A modification of the PCR assay along these lines might improve sensitivity.

Limited value of two widely used enzyme immunoassays for detection of Chlamydia trachomatis in women.

Enzyme immunoassays (EIAs) are widely used to diagnose chlamydial infections in patients attending genitourinary medicine clinics. They are relatively easy to perform and are suitable for testing large numbers of samples. The objective of this study was to determine what proportion of women with chlamydial infection, defined as the presence of Chlamydia trachomatis in a cervical smear or deposit and/or in the urinary tract, detected by means of a sensitive direct fluorescent antibody test could also be identified by using two commercially available EIAs to test cervical samples. On hundred fifty-one women attending the genitourinary medicine clinic at St. Mary's Hospital, London, were enrolled. The use of the Chlamydiazyme (Abbott Diagnostics, UK) and MicroTrak (Syva, UK) EIAs resulted in the identification of only 56% and 63%, respectively, of women with chlamydial infection detected by direct fluorescent antibody staining. Thus, the EIAs available for detection of chlamydiae in cervical samples are inadequate for identifying all infected women. Improvement might be achieved by testing multiple samples or by resorting to tests of greater sensitivity.

Chlamydia trachomatis in women: the more you look, the more you find.

OBJECTIVE: To determine the extent to which testing of multiple sites and samples is required to define whether a woman is Chlamydia trachomatis-positive. DESIGN: One-hundred and fifty women attending the Genitourinary Medicine clinic at St Mary's Hospital were enrolled; they had not received antichlamydial antibiotics in the previous three months, were not in a high-risk group for HIV infection, or pregnant, or using an intrauterine contraceptive device. Thirty-two women were re-examined three months after recruitment. METHODS: An urethral specimen was Gram stained (smear) and cultured for gonococci. Another urethral specimen was taken to detect C trachomatis elementary bodies (EBs) by the MicroTrak direct fluorescent antibody (DFA) test (Syva). An endocervical swab specimen was Gram stained (smear) and cultured for gonococci. One of two other endocervical swabs was used for the DFA test and was then placed in medium which was centrifuged in a MicroCentaur at 13,000 rpm for 10 min; the deposit was examined by using the DFA test. The first 15-20 ml of voided urine (first pass urine; FPU) was also centrifuged and the deposit tested similarly. RESULTS: Of 182 cervical smears and/or deposits tested for C trachomatis, 38 were positive; more cervical deposits (37) than smears (26) were positive and, of these, one-fifth of the deposits and one-third of the smears contained fewer than 10 elementary bodies. Of 162 paired urethral smears and FPU deposits available, one or other specimen of 36 pairs was chlamydia-positive, that is 31 smears and 32 deposits; of these, two-fifths of the smears and half of the deposits contained fewer than 10 EBs. Of 150 sets of cervical and urinary tract samples tested, 31 were chlamydia-positive at both sites, six in the cervix alone and four in the urinary tract alone. Of 139 women for whom there were valid first visit sample results, 36 (26%) were chlamydia-positive in the cervix, 34 (25%) in the urinary tract and 41 (29%) had at least one sample from either site positive. Overall, DFA tests of deposits from centrifuged cervical specimens achieved the highest sensitivity (88%) and those of cervical smears the lowest (70%). CONCLUSIONS: Deposits from centrifuged cervical specimens were C trachomatis-positive more often than were cervical smears. Testing deposits from centrifuged urines was as successful as testing urethral smears. One-fifth (cervical deposits) to one-half (urine deposits) of specimens contained fewer than 10 EBs. The urinary tract was chlamydia-positive almost as frequently as the cervix but both sites needed to be tested to define whether a woman was chlamydia-positive.

Regulation of prostaglandin endoperoxide H synthase by glucocorticoids and activators of protein kinase C in the human amnion.

Since glucocorticoids decrease and protein kinase C (PKC) activators increase amniotic PGE2 production, the possibility that they regulate the activity of prostaglandin endoperoxide H synthase (PGHS), the rate-limiting enzyme of prostaglandin synthesis from arachidonate, was investigated. Glucocorticoids inhibited the production of PGE2 from exogenous arachidonate specifically and in a concentration dependent fashion. Furthermore, cortisol decreased PGHS activity and the amount of PGHS protein in amnion microsomes, and reduced the rate of recovery of PGHS after acetylsalicylic acid (ASA) pretreatment. Actinomycin D blocked the inhibition of PGHS recovery by cortisol, but did not suppress the spontaneous recovery of the enzyme, indicating that the glucocorticoid induced a post-transcriptional inhibitor of PGHS synthesis. PKC-activating phorbol esters, such as 12-tetradecanoyl phorbol 13-acetate (TPA) increased the synthesis of PGE2 from exogenous arachidonate, also in a specific and concentration dependent manner. PGHS recovery after ASA treatment was enhanced by TPA. PGHS activity and protein concentrations were increased by phorbol ester treatment; however, this was apparent only in tissues in which the concentrations of PGHS were initially low. These results show that the synthesis of PGHS is positively and negatively regulated in the human amnion by PKC and glucocorticoids, respectively, and suggest that effectors using these pathways may regulate the enzyme in vivo.

Evaluation of sensitivity of 10 diagnostic assays for Chlamydia trachomatis by use of a simple laboratory procedure.

AIMS: To determine the sensitivity of commercially available diagnostic assays for Chlamydia trachomatis using a simple method. METHODS: Nine commercial assays and an "in-house" polymerase chain reaction (PCR) were evaluated using serial dilutions of a laboratory grown H serovar--four of them using a laboratory grown E serovar. Seven of the assays were further tested using dilutions of several cervical samples known to contain chlamydiae. RESULTS: The most sensitive assays were the MicroTrak direct fluorescent antibody (DFA) test (Syva) and the PCR which detected C trachomatis at a 10(-8) dilution of the H serovar, while the two least sensitive, Clearview (Unipath) and TestPack (Abbott), were positive only at 10(-4) and 10(-3) dilutions, respectively. A range of enzyme immunoassays (EIAs) and a nucleic acid hybridisation test were of intermediate sensitivity. The results with serovar E were consistent with these. When clinical samples were examined, the DFA test detected C trachomatis in dilutions at least 10-fold greater than any other assay. CONCLUSIONS: The range of sensitivity of diagnostic assays determined by the laboratory dilution procedure is very wide. Sensitivity assessed in this way, however, reflects the ability of the assays to detect C trachomatis in large scale clinical trials. The dilution procedure, which is simple to undertake, could therefore be applied by any laboratory before a new diagnostic method is considered for routine use.

Evaluation of sensitivity of 10 diagnostic assays for Chlamydia trachomatis by use of a simple laboratory procedure.

AIMS: To determine the sensitivity of commercially available diagnostic assays for Chlamydia trachomatis using a simple method. METHODS: Nine commercial assays and an "in-house" polymerase chain reaction (PCR) were evaluated using serial dilutions of a laboratory grown H serovar--four of them using a laboratory grown E serovar. Seven of the assays were further tested using dilutions of several cervical samples known to contain chlamydiae. RESULTS: The most sensitive assays were the MicroTrak direct fluorescent antibody (DFA) test (Syva) and the PCR which detected C trachomatis at a 10(-8) dilution of the H serovar, while the two least sensitive, Clearview (Unipath) and TestPack (Abbott), were positive only at 10(-4) and-3 dilutions, respectively. A range of enzyme immunoassays (EIAs) and a nucleic acid hybridisation test were of intermediate sensitivity. The results with serovar E were consistent with these. When clinical samples were examined, the DFA test detected C trachomatis in dilutions at least 10-fold greater than any other assay. CONCLUSIONS: The range of sensitivity of diagnostic assays determined by the laboratory dilution procedure is very wide. Sensitivity assessed in this way, however, reflects the ability of the assays to detect C trachomatis in large scale clinical trials. The dilution procedure, which is simple to undertake, could therefore be applied by any laboratory before a new diagnostic method is considered for routine use.