Development of an enzyme-linked immunosorbent assay for Bartonella henselae infection detection.

UNLABELLED: Several serological diagnostics rely on enzyme-linked immunosorbent assay (ELISA) to detect bacterial infections. However, for some pathogens, including Bartonella henselae, diagnosis still depends on manually intensive, time-consuming assays including micro-immunofluorescence, Western blotting or indirect immunofluorescence. For such pathogens, there is obviously still a need to identify antigens to establish a reliable, fast and high-throughput assay (Dupon et al. ). We evaluated two B. henselae proteins to develop a novel serological ELISA: a well-known antigen, the 17-kDa protein, and GroEL, identified during this study by a proteomic approach. When serum IgG were tested, the specificity and sensitivity were 76 and 65·7% for 17-kDa, respectively, and 82 and 42·9% for GroEL, respectively. IgM were found to be more sensitive and specific for both proteins: 17-kDa protein, specificity 86·2% and sensitivity 75%; GroEL, specificity 97·7% and sensitivity 45·3%. IgM antibodies were also measured in lymphoma patients and patients with Mycobacterium tuberculosis infection to assess the usefulness of our ELISA to distinguish them from B. henselae infected patients. The resulting specificities were 89·1 and 93·5% for 17-kDa protein and GroEL, respectively. Combining the results from the two tests, we obtained a sensitivity of 82·8% and a specificity of 83·9%. Our work described and validated a proteomic approach suitable to identify immunogenic proteins useful for developing a serological test of B. henselae infection. SIGNIFICANCE AND IMPACT OF THE STUDY: A reliable serological assay for the diagnosis of Cat Scratch Disease (CSD) - a pathological condition caused by Bartonella henselae infection - has not yet been developed. Such an assay would be extremely useful to discriminate between CSD and other pathologies with similar symptoms but different aetiologies, for example lymphoma or tuberculosis. We investigate the use of two B. henselae proteins - GroEL and 17-kDa - to develop a serological-based ELISA, showing promising results with the potential for further development as an effective tool for the differential diagnosing of B. henselae infection.

Epidemiological and molecular assessment of a measles outbreak in a highly vaccinated population of northeast Italy.

Two distinct measles outbreaks, unrelated from the epidemiological point of view but caused by genetically related strains, occurred in the Friuli Venezia Giulia region of northeastern Italy. Forty-two cases were reported during the period April-May 2008. In the first outbreak the index case was a teacher who introduced the virus into the Pordenone area, involving eight adolescents and young adults. The other concomitant outbreak occurred in the city of Trieste with 33 cases. The containment of the epidemics can be explained by the high MMR vaccine coverage in an area where the first dose was delivered to 93·4% and the second dose to 88·3% of the target children. Phylogenetic analysis of 14 measles virus strains showed that they belonged to a unique D4 genotype indistinguishable from the MVs/Enfield.GBR/14.07 strain, probably introduced from areas (i.e. Piedmont and Germany) where this genotype was present or had recently caused a large epidemic.

Epidemiology and effects of gastrointestinal nematode infection on milk productions of dairy ewes.

66 Pampinta breed ewes were studied during milking to evaluate the infection and the effect of gastrointestinal nematode on milk production sheep system. Naturally infected ewes on pasture were randomly allocated to two groups: TG, suppressively treated group every four weeks with levamisole and UG, untreated group. Faecal nematode egg counts and larval differentiation were conducted monthly. Successive groups of worm free tracer lambs were grazed with ewes and then slaughtered for worm counts. Test-day milk yield of individual ewes was recorded and ewe machine-milking period length (MPL) were estimated. Faecal egg counts and tracer nematode numbers increased towards midsummer and declined sharply toward the end of the study. TG (188.0 +/- 60 liters) produced more (p < 0.066) milk liters than UG (171.9 +/- 52.2) and TG had significantly more extended (p < 0.041) MPL than those of UG. The present study showed that dairy sheep were negatively affected by worms, even when exposed to short periods of high acute nematode (mainly Haemonchus contortus) infection.

Transmission of Burkholderia cepacia complex: evidence for new epidemic clones infecting cystic fibrosis patients in Italy.

To analyze national prevalence, genomovar distribution, and epidemiology of the Burkholderia cepacia complex in Italy, 225 putative B. cepacia complex isolates were obtained from 225 cystic fibrosis (CF) patients attending 18 CF centers. The genomovar status of these isolates was determined by a polyphasic approach, which included whole-cell protein electrophoresis and recA restriction fragment length polymorphism (RFLP) analysis. Two approaches were used to genotype B. cepacia complex isolates: BOX-PCR fingerprinting and pulsed-field gel electrophoresis (PFGE) of genomic macrorestriction fragments. A total of 208 (92%) of 225 isolates belonged to the B. cepacia complex, with Burkholderia cenocepacia as the most prevalent species (61.1%). Clones delineated by PFGE were predominantly linked to a single center; in contrast, BOX-PCR clones were composed of isolates collected either from the same center or from different CF centers and comprised multiple PFGE clusters. Three BOX-PCR clones appeared of special interest. One clone was composed of 17 B. cenocepacia isolates belonging to recA RFLP type H. These isolates were collected from six centers and represented three PFGE clusters. The presence of insertion sequence IS 1363 in all isolates and the comparison with PHDC reference isolates identified this clone as PHDC, an epidemic clone prominent in North American CF patients. The second clone included 22 isolates from eight centers and belonged to recA RFLP type AT. The genomovar status of strains with the latter RFLP type is not known. Most of these isolates belonged to four different PFGE clusters. Finally, a third clone comprised nine B. pyrrocinia isolates belonging to recA RFLP type Se 13. They represented three PFGE clusters and were collected in three CF centers.

Low rates of antimicrobial resistance in respiratory pathogens from a pediatric population in north-eastern Italy.

BACKGROUND: Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes are the main agents of bacterial upper respiratory tract infections in infants and children. In the past decades, the proportion of patients infected with drug-resistant organisms has increased; surveying at a regional level is important for understanding the size of the problem and elaborating therapeutic guidelines based on local epidemiology. METHODS: Since 1997, all isolates of Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes have been surveyed for antimicrobial resistance. RESULTS: In our zone, we found 3.5% penicillin-resistant and 18% macrolide-resistant Streptococcus pneumoniae, 15.8% ampicillin-resistant Haemophilus influenzae and 24.8% erythromycin-resistant Streptococcus pyogenes. CONCLUSIONS: Penicillin-resistant Streptococcus pneumoniae (PRSP) in our area does not represent a real concern; conversely, macrolide resistance in Streptococcus pneumoniae is relatively frequent. Resistance to ampicillin in Haemophilus influenzae in our area seems relatively low, due to beta-lactamase production. Even for Streptococcus pyogenes, macrolide resistance rates in our region are relatively low if compared with other countries and other italian regions. In conclusion, in our area penicillins are still the first choice drugs, at least in noncomplicated or not recurrent infections.

Case report: Salmonella panama osteomyelitis in a Ghanaian child with sickle cell disease.

Sickle cell disease is a rare condition in italian patients and even rarer are its complications, in particular Salmonella osteomyelitis. We describe a case of a Ghanaian child with sickle cell disease who developed osteomyelitis due to Salmonella panama, treated successfully with surgical debridement, followed by a prolonged period of specific antibiotic therapy.

Effect of infectious status and parity on somatic cell count and California mastitis test in pampinta dairy ewes.

The relationship between somatic cell counts (SCC) and California mastitis test (CMT) results according to the infectious status of mammary halves and parity of Pampinta dairy ewes was evaluated. Tests were associated to bacteriological analysis and classified into three groups: uninfected (negative culture), infected by minor pathogens and infected by major pathogens. Coagulase-negative Staphylococcus (32.4%), Micrococcus spp. (32.4%), Corynebacterium spp. (5.4%), and Bacillus spp. (1.4%) were the minor pathogens isolated, while Staphylococcus aureus (27%) and Escherichia coli (1.4%) were the major pathogens isolated. A good correlation was found between the CMT and SCC, which included inflammatory and epithelial cells (r = 0.64; P < 0.0001). SCC averages for the CMT scores shown in parentheses were 223 576 (0); 245,248 (1); 397,778 (2); 1,159,109 (3) and 2,460,833 (4) cells/ml. The correlation between SCC and the infectious status of udder halves was 0.58 (P < 0.0001). The relationship between SCC and CMT profiles and infectious status studied by a discriminant analysis showed, with an accuracy of 65%, three infectious status groups. SCC arithmetic means were 244,470 cells/ml for negative culture, 1,044,100 cells/ml for minor pathogens and 2,045,652 cells/ ml for major pathogens. With the exception of 1-year-old ewes, no significant differences were observed in SCC as affected by age or parity.

Effects of previous suppressive anthelmintic treatments on subsequent nematode infection in fattening cattle in Argentina.

The effect of previous suppressive anthelmintic treatments after weaning on parasitological parameters and weight gain of cattle was studied in the Pampeana region of Argentina. The study was carried out at two grazing fattening periods: April 1995/July 1996 and April 1997/July 1998. During both periods, 60 weaned calves that grazed contaminated pastures, were divided into three groups during the first part of the periods: GY1 group was treated every 2 weeks with doramectin while GY2 and GY3 groups remained untreated. During the second part of the periods, from October onwards GY1 and GY2 remained untreated and GY3 was treated every 2 weeks. In this second period two new groups of 20 weaning young calves were added: TG (treated every 2 weeks) and UG (untreated). Egg counts (EPG), larval cultures, pasture larval counts, serum pepsinogen (Pep) and live weight gain (LWG) were recorded monthly. Ostertagia, Cooperia, Trichostrongylus and Haemonchus were the predominant genera. Despite low levels of previous infection during the first part of the period, slight differences of EPG between GY1 (P<0.09) or UG (P<0.05) and GY2 were detected in the second part of the fattening period in 1995/1996. In 1997/1998 moderate infection levels during the first part of the period were observed. During the second part of this period, GY1 and UG showed higher (P<0.001) EPG than GY2, and only GY3 and TG had (P<0.05) lower Pep levels. Also, during the second part of 1997/1998, LWG responses of GY3 were higher (P<0.001) than those of GY1 and GY2. Live weight gain of GY2 exceeded GY1 by 10.7kg (P<0.006). Higher EPG and lower LWG of GY1 suggest that suppressive treatments negatively affected the level of resistance to infection of yearlings, but these effects were influenced by previous levels of nematode infection. The lack of differences between yearling (GY1) and calves (UG) groups suggest that, under the conditions of this study, there was no evidence that resistance to infection and the related parameters are influenced by the age.

[Gynecologic infections in adolescence]. / Le infezioni ginecologiche nell'adolescenza.

BACKGROUND: Adolescence in the age bracket with the highest incidence of sexually transmitted diseases (STDs). An observational study was performed in symptomatic patients attending our Outpatient Clinic for STDs in order to evaluate the diffusion of genital infections in the adolescent population and the different epidemiological distribution compared to adults. METHODS: The incidence of these infections was compared in two groups of patients attending the clinic between April 1995 and April 1999. The first consisted of 54 teenagers (13-19 years-old) and the second, used as a control, consisted of 917 women of child-bearing age (20-40 years). All patients underwent clinical examinations and microbiological tests of vaginal secretions (bacterioscopic test on coloured plates using the Gram method, cold tests, cultural assay of bacteria and fungi, assay for (Chlamydia trachomatis, Mycoplasma spp., Neisseria gonorrhoeae, Herpes simplex virus). RESULTS: The most common genital infection in teenagers was candidosis (35% vs 23.9%), followed by bacterial vaginosis (31.9% vs 25.8%) and mixed flora vaginitis (25.9% vs 22.6%). HSV was found in 1.8% of adolescents (vs 0.6%), in selected cases, Chlamydia and Mycoplasma were present in 16.6% (vs 1.1%) and 50% (vs 28%) of teenagers respectively. CONCLUSIONS: All genital infections were more frequent in the study population of adolescents compared to the control group. The adolescent population should be the target of a serious and systematic programme of prevention in order to ward off the sequelae of early and often irresponsible sexual activity.

Physiological and parasitological responses to nematode infections of fattening cattle in the Western Pampas of Argentina.

The epidemiology of nematode infection was studied in fattening grazing cattle from weaning (April 1994) to market at the end of their second autumn (July 1995). Sixty Aberdeen Angus calves of seven months of age were randomly allocated by weight to two groups: GT, treated every three weeks with doramectin (200 mcg/kg); and GI, an infected group, only treated with fenbendazole (7.5 mg/kg) at weaning and on the 1st of October. The two groups were grazed together on contaminated lucerne pastures until July, on 'clean' oat pastures until October and again on contaminated lucerne until the end of the trial. Fecal egg counts (epg), herbage larvae (L3), serum pepsinogen (Pep) and blood eosinophils (Eo) were evaluated monthly. Eight steers were slaughtered for worm recovery, three in July 1994, three in December 1994 and two in July 1995. Grazing feed intake was estimated by fecal output (chromic oxide method)/l-diet digestibility and to measure non-specific response, Brucella antibodies were detected at 11 and 40 days post-vaccination in early winter. Fecal egg counts, Pep and Eo of GI increased (P<0.01) from April to July when there was a moderate-to-high level of infection. Ostertagia, Trichostrongylus and Cooperia were the predominant genera. By late winter, all parameters decreased on oat 'clean' pastures and increased again when cattle returned to moderately infected lucerne. During summer, the parameters measured reflected the negligible numbers of L3 on pastures until early autumn. At this time, increased numbers of L3 were followed by a moderate rise (P<0.01) of epg, Pep and Eo values. During winter, GT showed higher (P<0.04) Brucella antibody IgG titers while feed intake of GI was 24.9% depressed (P<0.02). There were total cumulative weight-gain (WG) differences (P<0.001) between groups (GT=263.1 kg; GI=214.3 kg). During the second autumn, the mean WG of GT steers was 16.6 kg greater (P<0.04) than that of GI. Vaccination titres against Brucella suggested non-specific depression of immunity, while higher Pep and Eo levels in second-year steers may have reflected hypersensitivity reactions.

Risk factors for breast cancer mortality among the National Breast Screening Study of Canada participants.

We studied the relationship between risk factor information and breast cancer mortality by means of a case control study, nested within the population of the National Breast Screening Study of Canada (NBSS). The characteristics of women aged 40-59 years, identified at the initial screen, who subsequently died of breast cancer up to 7 years from the initial screen, were compared with those of controls drawn from the same population. Among the factors evaluated in this study, number of live births and presence of symptoms in the breast revealed on direct questioning were found to be significantly related to breast cancer mortality. The results suggest a decrease in risk of dying of breast cancer associated with one or more live births (OR: 0.79, 95% CI: 0.68, 0.93), and an increase in risk of dying of breast cancer associated with symptoms in the breast revealed on direct questioning at the initial screen (OR: 2.01, 95% CI: 1.20, 3.37).

Comparative effects of nematode infection on Bos taurus and Bos indicus crossbred calves grazing on Argentina's Western Pampas.

Breed differences in resistance or tolerance to naturally acquired gastrointestinal nematodes were compared between Aberdeen Angus (AA) and Santa Gertrudis (SG) weaned steer calves grazing in a temperate environment. Within breeds, one half of the calves was untreated: AAU (n = 17) and SGU (n = 18) while the other half was treated with moxidectin monthly: AAT (n = 14) and SGT (n = 14). All calves were grazed on the same contaminated pasture from autumn to winter, when each group was separated to uninfected paddocks for the remaining period of the study. Faecal egg counts, plasma pepsinogen levels (PPL), herbage larval counts and liveweight (LW) were recorded monthly. Egg counts and PPL of AAU and SGU increased from autumn to early winter when calves showed heavy parasitism (mainly Trichostrongylus axei) and severely sick calves needed emergency treatment. On the clean paddocks, parasitological parameters progressively decreased. Late winter egg counts were higher (P < 0.05) in SGU than in AAU. PPL of SGU were higher than AAU ones, but no significant differences were detected. Larval differential counts of SGU showed fewer Cooperia (P < 0.08) and higher T. axei (P < 0.002) proportions than those of AAU. Numbers of severely sick and dead SGU calves (61%) were greater (P < 0.01) than those numbers of AAU calves (17.5%). Cumulative LW gains differences within SG breed were higher (P < 0.0001) than those differences within AA breed (P < 0.003). LW gains of AAU during autumn-winter period were greater (P < 0.002) than those of SGU.(ABSTRACT TRUNCATED AT 250 WORDS)

The epidemiology of helminth infections of growing sheep in Argentina's western pampas.

Seasonal population trends of helminth parasites of growing lambs were investigated over 4 years. Successive worm-free lambs were grazed together with untreated lambs for 20-30 days and then slaughtered for helminth counts 2 weeks after their removal from pasture. Likewise untreated lambs from the same flock were slaughtered from 2 to 15 months of age for worm counts in the same way as tracer lambs. The predominant parasites were Haemonchus, Nematodirus and Trichostrongylus. Haemonchus contortus was found to be of major importance and the flock acquired massive worm burdens from summer to mid-autumn. Minimum burdens were seen from winter to early spring and maximum L4 stages were found from mid-autumn to early winter. Nematodirus burdens increased from December with a peak in late summer and then decreased to low values. Maximum larval availability was in autumn to early winter. Trichostrongylus (mainly T. colubriformis) populations increased in autumn and peaked in June-July, while the highest larval availability was in autumn. The minor genera recovered were Ostertagia, Cooperia, Trichuris, Oesophagostomum and Moniezia. Dictyocaulus, Chabertia and Teladorsagia were noted occasionally and liver flukes were not detected. No important and pathogenic numbers of L4 stages were seen and all predominant species were able to survive over summer or winter in pasture.

Fluoride-induced early teeth wearing in Argentinian cattle.

The relationship between observed early teeth wearing in cattle and the existence of large areas where excess fluoride in underground water prevails in Argentina, but has never been studied. Average values of bone fluoride exceeding 5000 ppm and well water containing up to 10.5 ppm of this element were found in a farm (ETWF) where early teeth wearing is observed, while in a control farm (NETWF), those values were 1480 ppm and 3.0 ppm respectively. Urine samples from animals from the ETWF contained almost 4 times more fluoride than those from the latter. Forage in both farms contained about the same levels of fluoride (15 to 25 ppm). None of the farms was exposed to industrial, volcanic, or geothermal activity. Lactating and adult animals were transferred from one farm to the other. After 30 months, adult animals from the NETWF showed no damage in teeth, while adults from the ETWF continued a rapid teeth wearing. Young animals from the NETWF developed severely damaged permanent teeth, while young animals from the ETWF developed normal teeth. We concluded that the problem exists in the ETWF and being a standard farm, the condition could be extrapolated to other areas.

Epidemiology and effects of nematode infections on beef cow-calf systems of Argentina's western pampas.

The epidemiology and the effects of nematode infections on cow-calf systems were followed from 3 weeks before calving to 7 months afterwards. Two groups, each of 15 cow-calf pairs grazed on separate lucerne pastures. Group I (GI) were treated monthly with oxfendazole (4.5 mg kg-1), cows being dosed from calving and calves starting 45 days later. Animals in Group II (GII) were not treated. The egg output of the cows was very low. An increase was recorded 2 months after parturition, consisting mainly of Ostertagia spp. The egg output and worm burdens of calves remained low until late summer and reached a peak in autumn. Ostertagia, Cooperia and Haemonchus were the main genera recovered from slaughtered calves. The pasture contamination and tracer calf worm counts remained consistently low until autumn when they began to increase. Inhibited early fourth stage larvae of Ostertagia were recovered during spring. After calving, the live-weight gains (LWG) of treated GI cows were significantly higher (P less than 0.004) than those of GII cows, whereas the LWG of GI calves were significantly higher during December (P less than 0.037) and March (P less than 0.029) than those of GII calves. There were significant (P less than 0.04) cumulative LWG responses between GI and GII calves with no differences in cow cumulative LWG at the end of the trial. For spring-born calves, these results suggest that strategic deworming programs in cow-calf systems may produce benefits.

Spiculopteragia spiculoptera, S. asymmetrica and Ostertagia leptospicularis from Cervus elaphus in La Pampa, Argentina.

The nematode species Spiculopteragia spiculoptera, Spiculopteragia asymmetrica and Ostertagia leptospicularis were recovered from the abomasum of wild red deer, Cervus elaphus, in the forest area of the semi-arid Pampeana Region of La Pampa (Argentina) for the first time. Oesophagostomum venulosum and Dictyocaulus spp. were also found.

Effects of an integrated control programme with ivermectin on growth, carcase composition and nematode infection of beef cattle in Argentina's western pampas.

The effect of previous pasture management and ivermectin treatment on gastrointestinal parasitism, weight gain and carcase composition of steers was studied in the semiarid Pampeana region of Argentina from April 1986 to April 1987. Three groups, each of 15 weaned calves, were grazed on separate lucerne pastures. Group 1 control calves, which grazed paddocks previously grazed by nematode-infected weaners and yearlings with a high nematode egg output, were only medicated when heavy parasitism was recorded. Group 2 calves, which grazed paddocks previously infected by steers at least two and a half years old with low nematode egg output, were treated strategically with ivermectin. Group 3 calves, which grazed on 'clean' paddocks, were treated monthly with ivermectin. Group 1 calves showed heavy parasitism and parasitological parameters were higher than in groups 2 and 3. The liveweight gain responses of groups 2 and 3 were significantly greater than those of group 1 (P less than 0.001) during autumn, winter and early spring. At the end of the study when cattle reached market condition, the liveweight gains of groups 2 and 3 were 74.1 and 81.9 kg, respectively, greater than group 1. Carcase analyses showed significantly greater weight and killing-out percentages in groups 2 and 3 than in group 1. Reduced total bone, muscle and fat weights were observed in group 1.

Epidemiology, effects and control of nematode infections on Zebu crossbred, Hereford and Hereford X Brahman calves of Argentina's western pampas.

Gastrointestinal parasitism of Zebu crossbred (Z), Hereford (H), and Hereford X Brahman (HB) weaned steer calves was observed from March 1985 to May 1986. Three groups of 60 calves were randomised and in each group the three breeds were equally present. Calves in GI were untreated, in GII were treated strategically and in GIII treated monthly with anthelmintic. The effects of anthelmintic treatments on the growth of cattle were assessed by comparing mean live-weight gain (MLG) responses of treated and untreated calves grazing the same permanent pasture. Worm recovery and egg counts increased from autumn to winter. After abundant July rainfalls, herbage infective larvae (L3) and worm burdens (mainly Ostertagia) reached the highest levels and Z calves presented symptoms of clinical parasitism. After high availability of L3 in spring, L3 and egg counts decreased to the end of the study. From the end of winter until summer a high percent of inhibited Ostertagia EL4 was seen. Blood samples showed reduced serum copper lower levels for GI. MLG responses of Z treated calves were significantly higher during the autumn-spring and summer-autumn periods. MLG responses of treated H calves were significant only during the winter-spring period. For HB calves there were no significant MLG responses during any period. The highest monthly treatment response was observed during winter and early spring. All strategic treatments gave significant MLG responses in spring and late summer. Initial effects of parasitism, from autumn until early spring had the greatest consequences in current and later productivity. For Z calves herbage L3, worm burdens, egg counts and MLG responses were higher than for H and HB calves. The Z breed were significantly heavier (+22.7%) at the end of the experiment in GIII than in GI. Similar trends were recorded, significantly, for H breed (+17.7%) and not statistically significant for HB (+12.7%). Under our temperate conditions it seems that H and HB calves showed greater capacity to resist nematode infection than Z calves.