Staphylococcus aureus adlb gene is associated with high prevalence of intramammary infection in dairy herds of northern Italy: A cross-sectional study.

Staphylococcus aureus is a major mastitis pathogen in dairy cattle worldwide, responsible for substantial economic losses. Environmental factors, milking routine, and good maintenance of milking equipment have been described as important factors to prevent intramammary infections (IMI). Staphylococcus aureus IMI can be widespread within the farm or the infection can be limited to few animals. Several studies have reported that Staph. aureus genotypes differ in their ability to spread within a herd. In particular, Staph. aureus belonging to ribosomal spacer PCR genotype B (GTB)/clonal complex 8 (CC8) is associated with high within-herd prevalence of IMI, whereas other genotypes are generally associated with individual cow disease. The adlb gene seems to be strictly related to Staph. aureus GTB/CC8, and is a potential marker of contagiousness. We investigated Staph. aureus IMI prevalence in 60 herds in northern Italy. In the same farms, we assessed specific indicators linked to milking management (e.g., teat condition score and udder hygiene score) and additional milking risk factors for IMI spread. Ribosomal spacer-PCR and adlb-targeted PCR were performed on 262 Staph. aureus isolates, of which 77 underwent multilocus sequence typing. In most of the herds (90%), a predominant genotype was identified, especially Staph. aureus CC8 (30%). In 19 of 60 herds, the predominant circulating Staph. aureus was adlb-positive and the observed IMI prevalence was relevant. Moreover, the adlb gene was detected only in genotypes of CC8 and CC97. Statistical analysis showed a strong association between the prevalence of Staph. aureus IMI, the specific CCs, and carriage of adlb, with the predominant circulating CC and presence of the gene alone explaining the total variation. Interestingly, the difference in the odds ratio obtained in the models for CC8 and CC97 suggests that it is carriage of the adlb gene, rather than the circulation of these CCs per se, that leads to higher within-herd prevalence of Staph. aureus. In addition, the model showed that environmental and milking management factors had no or minimal effect on Staph. aureus IMI prevalence. In conclusion, the circulation of adlb-positive Staph. aureus strains within a herd has a strong effect on the prevalence of IMI. Thus, adlb can be proposed as a genetic marker of contagiousness for Staph. aureus IMI in cattle. However, further analyses using whole-genome sequencing are required to understand the role of genes other than adlb that may be involved in the mechanisms of contagiousness of Staph. aureus strains associated with high prevalence of IMI.

Short communication: Characterization of Staphylococcus aureus from bulk tank milk of dairy cattle in Lombardy (northern Italy).

Staphylococcus aureus is one of the most important pathogens causing mastitis in cattle, and it is responsible for economic losses in dairy herds worldwide. The PCR amplification of the 16S-23S rRNA intergenic spacer (ribosomal spacer PCR, RS-PCR) allows a rapid classification of the strains in genotypes and genotypic clusters (CL), which are characterized by different epidemiological and clinical properties. Both RS-PCR and multi-locus sequence typing (MLST) were performed on strains isolated from bovine bulk tank milk (BTM) collected from dairy herds located in the Lombardy region (northern Italy), to outline the distribution of Staph. aureus genotypes in this geographical area. Out of 844 examined samples, 398 were positive for Staph. aureus, with a variable count (cfu/mL) Up to 8 colonies from each sample were genotyped. A total of 1,101 Staph. aureus strains were analyzed with RS-PCR, and only a selection of them (n = 86), in relation to their frequency and geographical origin, underwent MLST. This study revealed 8 major genotypic clusters (CLB, CLC, CLR, CLS, CLI, CLF, CLAO, and CLZ), of which Staph. aureus CLB (29.3%) was the most common. Samples of BTM positive for CLB had a Staph. aureus cfu/mL count significantly higher than the non-CLB positive ones. Our MLST analysis showed genotypes already known as bovine-associated in literature, such as clonal complexes CC8, CC97, and CC151. The same selection of 86 strains was also analyzed for the presence of the adlb gene, which was recently proposed as a possible marker of contagiousness. Most Staph. aureus belonging to CLB or CC8 carried the adlb gene (85%), whereas this gene was detected in only 9% of non-CLB strains (CLAA, CLBI, CLBJ, CLS). In conclusion, the present study confirms that Staph. aureus CLB, which is recognized as a contagious genotype, is a particularly relevant agent of intramammary infection in dairy cows in Lombardy, and indirectly supports the idea that adlb can be a possible marker of contagiousness of isolates.

Animal-based measures on fattening heavy pigs at the slaughterhouse and the association with animal welfare at the farm level: a preliminary study.

Monitoring animal welfare (AW) in pig farms requires both proper indicators and a feasible approach. Animal-based measures (ABMs) are well-established AW indicators. Furthermore, AW screening at the slaughterhouses could be useful for identifying problems on farm. The aim of this study was to evaluate ABMs at the slaughterhouse and, when possible, to compare these ABMs with those collected on the farm. The study was carried out in northern Italy in a commercial abattoir and in a sample of farms. Animal-based measures were recorded on pigs from 62 batches of 54 farms, during ante-mortem (n=10 085 pigs) and post-mortem (n=7952 pigs) inspections. Sixteen of 54 farms were selected to compare ABMs collected at the slaughterhouse with ABMs collected on the farm. Overall, 2295 pigs (mean pigs examined per farm 119±45) were inspected at the slaughterhouse (group S) and 420 pigs (mean pigs per farm 26±5) on the farm (group F). Non-animal-based measures were also collected at the 16 farms. Differences between groups S and F, at the animal level, were assessed by a two-tailed paired Wilcoxon-Mann-Whitney test. Differences at the site of observation level (farm and slaughterhouse) were assessed by Fisher's exact test using a hierarchical log-linear modelling for contingency tables. The most frequent ABMs at the slaughterhouse were manure on the body (47.7%), followed by dermatitis (28.0%), white spot (25.4%) and bursitis (24.7%). Recording ABMs at the slaughterhouse and on the farm usually yielded similar results; however, there were some exceptions. In particular, significant differences were found for non-uniformity of size (P<0.05) and dermatitis (P<0.001), which were higher at the slaughterhouse than on the farm. Results of log-linear modelling underlined the effect of the farm of origin on the percentage of pigs with bursitis, manure on the body and ear injuries that were observed at the slaughterhouse. In group S, significant associations between manure on the body and insufficient presence of clean and dry areas in the corresponding farm were found (P<0.05). Although these results should be interpreted with care due to the limited sample of farms, the slaughterhouse could be a feasible site of observation of ABMs, which could then be integrated in monitoring of AW on farm. Considering the number of slaughtered batches per farm, it would be possible to repeat assessments several times throughout the year for each farm, which could help provide an index for the continuous monitoring of AW.

Prevalence of Staphylococcus aureus and of methicillin-resistant S. aureus clonal complexes in bulk tank milk from dairy cattle herds in Lombardy Region (Northern Italy).

Staphylococcus aureus is the most important causative agent of subclinical mastitis in cattle resulting in reduced milk production and quality. Methicillin-resistant S. aureus (MRSA) strains has a clear zoonotic relevance, especially in the case of occupational exposure. The aim of the study was to evaluate the prevalence of S. aureus and MRSA in bulk tank milk (BTM) from dairy cattle herds in the Lombardy Region (Northern Italy) and to identify the main MRSA circulating genotypes. MRSA strains were characterized by susceptibility testing, multi-locus sequence typing (MLST), spa typing and SCCmec typing. A total 844 BTM samples were analysed and S. aureus and MRSA were detected in 47·2% and 3·8% of dairy herds, respectively. MLST showed that the majority (28/32) of isolates belonged to the typical livestock-associated lineages: ST398, ST97 and ST1. Interestingly, in this study we report for the first time the new ST3211, a single locus variant of ST(CC)22, with the newly described 462 aroE allele. Our study indicates high diffusion of S. aureus mastitis and low, but not negligible, prevalence of MRSA in the considered area, suggesting the need for planning specific control programmes for bovine mastitis caused by S. aureus, especially when MRSA is implicated.

Genomic characteristics of Staphylococcus aureus strains associated with high within-herd prevalence of intramammary infections in dairy cows.

Staphylococcus aureus is one of the most important causes of mastitis in dairy cattle. Based on previous research, Staph. aureus genotypes with different pathogenic and contagious properties can cause intramammary infection (IMI) and coexist in the same herd. Our study aimed to compare Staph. aureus strains from herds that differed in IMI prevalence using different molecular approaches such as ribosomal spacer (RS)-PCR, multilocus sequence typing (MLST), spa typing, ribotyping, pulsed-field gel electrophoresis (PFGE), and multiplex PCR. For this purpose, 31 dairy herds with Staph. aureus IMI were selected, and 16 of these were chosen for a comparison study: the 8 high-prevalence (HP) herds had Staph. aureus IMI prevalence >28% and the 8 low-prevalence (LP) herds had an IMI prevalence <4%. A total of 650 isolates of Staph. aureus from mammary quarters of all positive cows were genotyped with RS-PCR, a technique based on amplification of a portion of the intergenic spacer 16S-23S rRNA, and a subset of 54 strains was also analyzed by multiplex PCR, ribotyping, PFGE, MLST, and spa typing. The RS-PCR analysis revealed 12 different profiles. Staphylococcus aureus strains isolated from 5 out of 8 HP herds showed a profile identical to the genotype B (GTB), described in previous studies as being strongly associated with high within-herd prevalence of Staph. aureus mastitis and the presence of the genes coding for enterotoxins sea, sed, and sej, a long x-region of spa gene, and 3 lukE fragments. Moreover, all strains isolated in the HP herds possessed genes coding for staphylococcal enterotoxins. In LP herds, a limited number of strains of 6 genotypes, different from those isolated in HP herds, were identified and GTB was not found. Within these genotypes, 4 strains were positive for the mecA gene. Preliminary results and comparison with other genotyping methods confirmed that genotyping by RS-PCR is an accurate, rapid, and inexpensive tool for future field studies on Staph. aureus mastitis strains and generates clinically relevant results.

Short communication: Prevalence of Staphylococcus aureus and methicillin-resistant S. aureus in bulk tank milk from dairy goat farms in Northern Italy.

Staphylococcus aureus is regarded as a leading cause of mastitis in goats. However, few data are available on the presence of methicillin-resistant S. aureus (MRSA) in this species. The aim of this study was to assess the prevalence of S. aureus and MRSA in bulk tank milk samples from dairy goat farms in Northern Italy. Eighty-five out of 197 samples (43.1%) tested positive for S. aureus with counts ranging from 10 to more than 1.5 × 10(4) cfu/mL. The MRSA was screened by both direct plating followed by a disk diffusion test to evaluate methicillin resistance and a selective enrichment method. Methicillin-resistance was confirmed by mecA-specific PCR. Methicillin-resistant S. aureus was identified in 4 samples (2.0%) and multilocus sequence typing (MLST) showed the presence of livestock-associated MRSA belonging to lineages ST398 (n = 3) and ST1 (n = 1). In one case we demonstrated that the same MRSA strain was able to persist over time on the farm, being isolated from both bulk tank milk and the udder of 3 goats 1 yr after the first isolation. The high prevalence of S. aureus-positive herds detected in this study and the presence of MRSA strains belonging to livestock-associated genotypes is of concern, and represents a novel finding in the Italian dairy goat production system. The application of stringent measures for the control of S. aureus mastitis at the farm level seems appropriate to reduce the economic losses, and to minimize the risk of foodborne illness and the transmission of MRSA to humans by occupational exposure.

Immunohistochemical detection of aetiological agents of proliferative and necrotizing pneumonia in italian pigs.

Proliferative and necrotizing pneumonia (PNP) is a form of interstitial pneumonia that occurs in weaning and post-weaning pigs. PNP is characterized by hypertrophy and hyperplasia of type II pneumocytes and coagulative necrosis and granular debris within alveolar spaces. Canadian and European studies suggest that the porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus type 2 (PCV2) are the main causes of the disease, but Aujezsky's disease virus (ADV) and swine influenza virus (SIV) have also been considered as potential aetiological agents. An immunohistochemical study was carried out on the lungs of 28 Italian pigs with PNP in order to evaluate the role of PRRSV, PCV2 and ADV in PNP lesions. PRRSV infection was identified in the lungs of 11 pigs, PCV2 in the lungs of four pigs and coinfection with both viruses in the lungs of eight pigs. Neither virus was detected in the lungs of the remaining five pigs. ADV antigen was not detected in any sample. The principle aetiological agent of PNP in Italy therefore appears to be PRRSV. Coinfection with PRRSV and PCV2 is characterized by more severe microscopical changes in affected lungs.

Application of a protocol for the diagnosis of postweaning multisystemic wasting syndrome in Italy.

Samples of superficial inguinal and bronchial lymph nodes, ileum, tonsil and lung were taken from three to five pigs on each of 61 farms with a clinical history of postweaning multisystemic wasting syndrome (PMWS). The samples were examined histologically and by immunohistochemistry for porcine circovirus type 2 (PCV-2). PMWS was diagnosed in two stages: first, an evaluation of the haematoxylin and eosin-stained sections that identified the cases in which the characteristic PCV-2 cytoplasmic inclusion bodies were apparent, and secondly, a conclusive step in which immunohistochemistry was applied to confirm PMWS in the cases in which there were positive immunohistochemical results that coincided with lesions indicative of PMWS in at least one of the lymphoid and/or lung tissues. The location of PCV-2 in specific lesions (cell depletion in lymphoid organs and interstitial pneumonia) confirmed PMWS in 45 of the 61 farms, 31 of which were also infected with porcine reproductive and respiratory syndrome virus. The lymphoid tissues were more reliable than the lungs for the diagnosis of PMWS, both in individual pigs and in groups of pigs, and farm diagnoses based on a group of pigs were more reliable than diagnoses based on single pigs.

Capillaria hepatica infection in wild brown rats (Rattus norvegicus) from the urban area of Milan, Italy.

Forty-seven wild brown rats (Rattus norvegicus) collected from the urban area of Milan (Italy) were screened for Capillaria hepatica liver infection. The liver of each rat was grossly and histologically examined for the presence of C. hepatica adults, eggs and typical C. hepatica induced lesions. In 17 rats (36%) liver lesions consistent with C. hepatica infection were detected. Grossly, white-yellow nodules of 1-5 mm in diameter were present, either scattered on the liver surface or localized in a single lobe. Histologically, granulomatous liver lesions associated with eggs and/or worms were observed. The degree of gross liver involvement was moderate in most of the positive cases (71%). About 30 cases of C. hepatica infection in humans have been documented world-wide, most of which are reported in children from 1 to 5 years of age. Our results suggest that the potential transmission of C. hepatica to children in the study area should be considered an important health issue.

Prevalence of pneumonia due to Legionella pneumophila and Mycoplasma pneumoniae in a population admitted to a department of internal medicine.

We studied 177 patients with pneumonia admitted to an internal medicine department over a period of 3 years to determine the incidence of two emerging pathogens, Legionella pneumophila and Mycoplasma pneumoniae. Clinical, radiological and laboratory tests were performed and included blood cultures, serology, gram staining and sputum cultures. L. pneumophila was the agent involved in 9 patients (5.1%) and M. pneumoniae in 12 (6.8%). These prevalences were about in the middle of the range of previously published figures. Legionella pneumonia is a rare illness, which even in the absence of suggestive clinical signs must be considered because of its possibly serious course and to allow appropriate therapeutic decisions to be made.

[Comparison between a diet with mainly vegetal protein content and a diet with mainly animal protein content in hepatic cirrhosis]. / Confronto tra una dieta a prevalente contenuto proteico vegetale ed una a prevalente contenuto proteico animale nella cirrosi epatica.

Eight cirrhotic patients were studied in a randomized crossover single blind study in order to compare the effects of two isocaloric, and with equal amount of protein diets on nitrogen balance and on the portal systemic index. One diet was 2:3 vegetable protein to total protein content (diet V) and the other 2:3 animal protein to total protein content (diet A). Each diet was changed every three weeks. Five patients followed the VAV scheme and three the AVA scheme. The mean values of the porto systemic index were almost the same with the two diets while the value of the nitrogen balance was less negative with the diet A than with the diet V, showing an increasing trend, more marked with the diet A, at every consecutive time of observation. The variance analysis was not significant for any of the two diets. Diet V was well tolerated.

Diagnostic outpatient aspiration curettage. Two years experience.

Endometrial aspiration curettage was performed on 1001 women, without anaesthesia. Our experience confirms that Vabra-curettage is, for patients, a simple and well tolerated sampling technic. We were not able to perform the technic in 18 patients for close cervical stenosis. In 41 patients the samples were quantitatively not adequate for a correct histological diagnosis. In all these cases a control by D & C was performed, in general anaesthesia, and no proliferative pathology was found. The most common presenting symptom in Intraepithelial Carcinoma of Endometrium (ICE) and in carcinoma was abnormal vaginal bleeding; nevertheless 5 cases of ICE and 2 cases of adenocarcinoma did not show any symptom. This study confirms that systematic screening of asymptomatic perimenopausal and postmenopausal women may lead to the discovery of occult endometrial cancers, some of which are advanced.

Respiratory function 8 years after a diagnosis of peripheral airways disease.

A program of screening for airways obstruction was designed to examine: (1) the precision and the reliability of the proposed tests (2) the evolution of respiratory impairment in 69 subjects, 8 years after a diagnosis of peripheral (small) airways obstruction. The tests that exhibit very good reliability are: vital capacity, total lung capacity, 1-second forced expiratory volume, single breath CO diffusion capacity, curvilinearity score; and the ratio of closing capacity to total lung capacity. 39% of smokers and 15% of nonsmokers developed central airways obstruction 8 years after the diagnosis of peripheral airways disease. These results confirm the hypothesis that small airways may be regarded as a noisy rather than a silent zone.

Small airways disease: a trial of an easy functional discrimination of preclinical emphysema.

In the genesis of small airways obstruction (SAO) we have to consider two alterations with different prognoses: the first, in which the obstruction depends on a reduction of the diameters of the small bronchi (i.e. with prevalent 'intrinsic mechanism)'), is reversible; the second, probably irreversible, is due to pulmonary emphysema in its pre-clinical stage. It is useful to distinguish between these two mechanisms by means of functional tests which are easily applicable in epidemiological screening. We have studied, by means of different lung mechanics tests, 5 normal, male, non-smoking subjects (N) and 10 male smokers considered to be affected by SAO. Several theoretical considerations, experimental data and prospective studies suggest that under various tests of easy execution for a screening program there are some which have, in some cases, a prognostic value and seem to be coupled to a qualitative meaning: this is the case for delta N2 which is significantly greater in subjects of the extrinsic group where the abnormality is considered irreversible. Some other tests are very sensitive to detect SAO subjects by they are lacking in specificity and in prognostic meaning (CV, VEmax, FDC). Finally, other tests [maximal mid-expiratory flow (MMEF) and residual volume (RV)] seem to have a prognostic value only because they indicate an advanced stage of the illness, not depending on the kind of the affection (SAO Ex or SAO In).

Peripheral pulmonary wedge angiography in chronic obstructive pulmonary disease. Relationship to pulmonary function, chest x-ray film, and hemodynamic data.

Peripheral pulmonary wedge angiographic studies can investigate only limited areas of the pulmonary vascular bed; nevertheless, this technique is very useful in emphasizing the morphologic changes of small vessels (less than 1 mm in diameter) in pulmonary emphysema. The aim of this work is to determine the relationship between peripheral wedge angiographic appearances and pulmonary function, the chest x-ray film, and hemodynamic data in patients with chronic obstructive pulmonary disease. Three healthy subjects, four patients with type-A chronic obstructive pulmonary disease, and eight patients with type-B chronic obstructive pulmonary disease were studied. Among the tests of pulmonary function, only a marked reduction (less than 60 percent of predicted) of the diffusing capacity for carbon monoxide is always indicative of severe widespread emphysema. No relationship was found between hemodynamic data and peripheral wedge angiographic findings. Some agreement resulted between the chest x-ray film and peripheral wedge angiographic abnormalities only in patients with type-A chronic obstructive pulmonary disease; on the contrary, in patients with type-B chronic obstructive pulmonary disease, both false-positives and false-negatives were noted.

Prospective study of airway obstruction in a population with small airway disease.

In this paper, we discuss results showing an increase of bronchial obstruction due to exposure in an iron foundry. The research has been conducted in two steps: in the first, we selected a sample of 100 subjects, all working in the iron foundry, who were affected only by small airway obstruction. In a second step, 30 months later, 99 of these subjects were re-examined and the present airway condition determined. In 43 subjects we found abnormal results of the tests indicating total airway obstruction after the 30 months. After properly accounting for ageing, we found that even in the subsample of non-smokers a deterioration had occurred, showing an effective contribution of environmental pollution. The use of RV, MMEF, CV, DLCO for detecting small airway obstruction is evaluated. The combination of RV, MMEF and CV is the most efficient mosaic of tests for the selection of subjects with small airway obstruction (89%) and with high probability of developing total airway obstruction (44%).

Drive and performance of ventilatory system during rebreathing.

In eight normal subjects, eight patients with asthma, and six patients with chronic obstructive lung disease, the first derivative of occlusion pressure at FRC during rebreathing has been studied. The maximal dP/dt was always lower in patients. Such a behavior may depend on reduced neuronal discharge as well as on reduced effectiveness of the contraction of the respiratory muscles. Though (dP/dt)max. may provide a reliable measure of the sensitivity of the respiratory center only in a homogeneous group, the results of plotting its value against ventilation and end-tidal Pco2 confirm the failure of the thoracomuscular system to convert the output of respiratory centers into increase of pulmonary pressure and ventilation.