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Microbiome and resistome successions in pig carcasses and fresh pork meat throughout slaughtering, processing and shelf-life.

BACKGROUND: Slaughterhouses and meat cutting plants represent potential hotspots for the spread and transfer of spoilage and pathogenic, including antimicrobial resistant, bacteria to meat and meat products. Here, we characterise the progression of the microbiome and resistome of two pork cuts (loin and sirloin) at different stages of processing, from the slaughter line to the end of shelf-life. To this end, we analysed samples from facility surfaces, carcasses, and meat cuts using whole metagenome sequencing. RESULTS: The taxonomic and antimicrobial resistance gene (ARG) profiles of carcasses and meat cuts were significantly influenced by the point of sampling and the processing room. The facility surfaces were found to be the main source of some abundant genera, such as Anoxybacillus, Acinetobacter, Pseudomonas, and Brochothrix, in carcasses and meat cuts. A total of 1,291 metagenome-assembled genomes were reconstructed, corresponding to the most prevalent species identified in the taxonomic analysis at the read level. A reduction in bacterial and ARGs richness and diversity was observed for carcasses and meat cuts along the production chain, which suggests that processing procedures are effective in reducing bacterial and ARGs loads. Nonetheless, an increase in the ARGs load was observed at two sampling points: the carcass after evisceration and the sirloin at the end of its shelf-life (in this case linked to the increase of a single gene, tet(L)). The ARGs most frequently detected were those associated with resistance to tetracyclines, aminoglycosides, and lincosamides. Acinetobacter (in processing environments and carcass/meat samples) and Staphylococcus (in carcasses and meat) were identified as the main genera associated with the ARGs found. CONCLUSIONS: Overall, our results provide the most detailed metagenomics-based perspective on the microbial successions of pig carcasses and fresh meat cuts during slaughtering, processing, and commercialisation. The observations made suggest that selection pressures imposed by processing steps and contact with facility surfaces contribute to shaping the microbiome and resistome of the two pork products throughout their production line and shelf-life. Video Abstract.

Animals

[Preservation of vitamin B1 and B2 during the preparation of pork meat].

Thiamin and riboflavin seem to resist fairly well to the proceedings of meat products studied here (cooked hams, patties, dry sausages). This result is normal enough to riboflavin, it is less for thiamin which sensitive to nitrite and to heating is well established by the literature. The origin of these observed phenomenons of protection is probably due to the complexation of these vitamins by the meat proteins. A similar phenomenon has been seen before by several authors on milk proteins, eggs and specially sulfhydril groups. This complexation is confirmed by the important release of both vitamins we noted during processing. They are determined much more easily on processed meat products than on raw materials. The nutritional repercussion of the observed phenomenon is difficult to estimate accurately. The vitamins release we find out, make them more available for the organism but it would be compared to the enzymatic decomplexation which occurs in vivo, during digestion, that we did not studied. Nevertheless such an approach would be necessary for the estimation of the practical value of the analytical determinations generally used.

Animals

Elucidating the evolution of meat quality, water distribution, microstructure, and protein structure during sous-vide and micro-pressure cooking.

This study investigated the evolution of eating quality (colour, texture and volatile flavour compounds), water status, microstructure and protein structure of pork meat under different cooking methods. The methods analysed included traditional cooking (TC: 10, 20, 30 and 40 min, 100 °C), sous-vide cooking (SV: 1, 2, 3 and 4 h, 60 °C) and micro-pressure cooking (MC: 10, 20, 30 and 40 min, 120 °C). Across the three cooking processes, as cooking time increased, cooking loss, lightness, yellowness, P23, β-sheet, random coil and surface hydrophobicity of the meat samples increased. By contrast, redness, P22, hydrogen proton density, esters content, α-helix, β-turn and sulfhydryl group content decreased. Moreover, the Warner-Bratzler shear force (WBSF), adhesiveness, hardness, springiness, gumminess, chewiness, alcohols, aldehydes, ketones and fluorescence intensity of the meat samples, initially increased and then decreased as cooking progressed. SV resulted in higher water-holding capacity (WHC), improved redness and increased alcohol and ester levels, whereas MC produced softer meat and greater water mobility. Furthermore, MC enhanced the degree of microstructural damage and protein structural unfolding in the meat. MC requires less time to achieve textures and flavours similar to those obtained using the TC and SV methods. Thus, MC is an efficient cooking method for the catering industry to obtain desired meat quality rapidly.

Cooking

[Data on the quantity and quality of lipolytic bacteria in products with a meat base].

The numbers of lipolytic bacteria and their proportion, relatively to total mesophilic aerobic bacteria, were determined in 65 samples. Other analyses: psychrotrophic bacteria, coliform bacteria, Escherichia coli, fecal streptococci, and enterotoxinogenic staphylococci were also made. The less modified products generally contained a low number of lipolytic bacteria. 101 isolates very lipolytic were obtained, 81 of which were Gram + bacteria and 20 were Gram --. It is hypothesized that in pork-meat preparations, Gram + bacteria, especially Bacillus and fecal steptococci, have a lipolytic action of greater importance than it is generally assumed.

Bacteria

Convergence and global molecular epidemiology of Klebsiella pneumoniae plasmids harbouring the iuc3 virulence locus: a population genomic analysis.

BACKGROUND: Klebsiella pneumoniae is an important pathogen of humans and animals. In the past five years, increasing reports of convergent strains that carry both virulence factors and antimicrobial resistance genes (ARGs) have raised serious public health concerns. The aim of this study is to describe the global diversity of plasmids carrying iuc3 (a key virulence factor in K pneumoniae associated with pigs and clinical isolates) from diverse settings, and their role in the emergence of convergent strains through hybridisation with plasmids carrying ARGs. METHODS: This population genomic analysis study was designed to describe both the global and local diversity of iuc3-carrying plasmids from diverse sources, and the co-occurrence of iuc3 with ARGs. We used all 4148 Klebsiella spp isolates from two large One-Health studies (SpARK, Italy, and OH-DART, Thailand), including 191 Klebsiella isolates from pigs, 635 from clinical isolates, 1040 from hospital and community carriage, and 2282 from other sources. Short-read sequencing of Klebsiella isolates was performed as part of the SpARK study. We sequenced Klebsiella isolates from the OH-DART (MicrobesNG, Birmingham, UK; HiSeq and NovaSeq, Illumina San Diego, CA, USA; GridION, Oxford Nanopore Technologies, Oxford, UK) and SpARK (MinION or GridION, Oxford Nanopore Technologies, Oxford, UK) studies. We also retrieved plasmid sequences carrying iuc3 from the National Centre for Biotechnology Information (NCBI). To ascertain the degree of diversity, evolutionary dynamics, and structuring across ecological and geographical axes, we detected ARGs and virulence loci, analysed clustering patterns and generated approximate maximum-likelihood phylogenetic trees. FINDINGS: We identified 48 K pneumoniae isolates with iuc3 in the SpARK data and 79 in the OH-DART data. Three (2·4%) of these 127 isolates were from clinical sources, 73 (57·5%) were from pig or pork meat. iuc3 isolates corresponded to multiple (n=47) host sequence types (STs), with ST35, ST45, ST881, ST25, and ST967 harbouring iuc3 in both datasets. We generated hybrid assemblies for 44 (SpARK) and 36 (OH-DART) isolates, plus a single iuc3 isolate from Germany. 53 (65·4%) of these isolates were from pigs, three (3·7%) from clinical sources, and 25 (30·9%) from other sources. There were an additional 48 iuc3 positive isolates from our collections for which only short read data was available. A single iuc3-positive Klebsiella oxytoca isolate from a pig farm was detected in the SpARK data, which was also sequenced. We identified 330 iuc3-positive isolates and 58 iuc3-carrying plasmid assemblies from NCBI, of which 83 (21·4%) were from clinical sources, 120 from pigs (30·9%), and 185 (47·7%) from other sources or of unknown provenance. These isolates were from K pneumoniae except two isolates of Klebsiella quasipneumoniae subsp similipneumoniae and one of Enterobacter hormaechei. The combined dataset of 517 iuc3 plasmids ranged in size from 110 375 bp to 365 580 bp and mostly corresponded to multiple IncFIB(K) and IncFII replicon types. We found seven convergent K pneumoniae plasmids in the Thai data: six from fresh markets and one from a neighbouring hospital. These plasmids emerged through the hybridisation of cocirculating iuc3 plasmids and plasmids encoding extended-spectrum β-lactamases (ESBLs), although none of these seven plasmids carried genes encoding carbapenemases. We also identified putative cocirculating parental plasmids carrying iuc3 and ESBL-encoding genes. Clustering and phylogenetic analysis resolved the iuc3 plasmid sequences into three groups, which were consistent using both complete plasmid sequences (n=139) and short-read data (n=517). In the complete plasmid sequence data, 66 strains contained group 1 plasmids, 38 strains contained group 2 plasmids, and 35 strains contained group 3 plasmids. Group 3 plasmids are mostly carried by isolates circulating in hospitals throughout Asia, with occasional examples in Europe and elsewhere, and carry multiple ARGs and potential virulence factors. By contrast, group 1 plasmids are commonly carried by porcine isolates in Europe, and group 2 are a heterogeneous mixture of geographical and ecological sources. INTERPRETATION: Plasmid hybridisation occurs frequently outside of the health-care environment and can lead to the convergence of resistance and virulence traits. Generating complete plasmid sequences from regional population-scale samples facilitates the identification of convergent plasmids and their putative parental plasmids. Three robust groups of iuc3 plasmids were resolved, which show both epidemiological and geographical differences; one of these groups was associated with clinical isolates in Asia and warrants targeted plasmid surveillance. FUNDING: UKRI, JPIAMR, Evolution Education Trust, and a Schlumberger Foundation Fellowship.

Plasmids

[Human trichinosis. A recent epidemic in Vientiane (Laos) (apropos of 32 cases)].

Fifty patients were hospitalised at Vientiane during May and June 1975. Infection was caused by consumption of pork meat (som-mou, lap mou and lap leuat). After an incubation period of 8 to 11 days intermittent diarrhoea followed by constant fever and orbital oedema occurred. Several days later diffuse myalgias occurred. Three patients presented neurological symptoms: left hemiplegia flexible paraplegia and limb paresis; these symptoms regressed spontaneously. 87% of the clinically suspected cases had specific serum antibodies (IFI, ID, IE). Hyperleucocytosis and hypereosinophilia were a constant factor. Anti-AH antibodies were detected by the Widal test. The histopathology characteristic of myositis was seen in the 12 muscle biopsies. Also observed was the precystic state of the larvae which is in agreement with the early timing of the samples which were obtained during the 3rd and 4rd week of infection.

Humans

The isolation of salmonellas from British pork sausages and sausage meat.

Between 1969 and 1974, 1467 packets (3309 samples) of pork sausages and sausage meat produced by two large and two medium sized manufacturers and several local butchers were examined for the presence of salmonellas. Of these, 435 packets (786 samples) were found to contain salmonellas, but there was a wide variation in the isolation rates according to the producer. The salmonella incidence in samples from several small and two medium sized producers was low (0-11%) while the results from the two large producers investigated showed a striking difference, the rate of salmonella contamination in the product of one was low (about 2%) and in that of the other consistently high (40-60%). A comparison of liquid enrichment media, incubation temperatures and selective agar media was also carried out to determine the most efficient combination for the isolation of salmonellas from minced meat products. The results showed that (a) incubation of enrichment cultures at 43 degrees C. yielded a consistently greater number of salmonella isolations that at 37 degrees C., regardless of plating medium, (b) tetrathionate broth A (Rolfe) was superior to selenite broth as en enrichment medium at both 37 and 43 degrees C. and (c) brilliant green agar gave better results than deoxycholate citrate sucrose agar and bismuth sulphite agar as a selective medium.

Animals

[Distribution of Cl. perfringens in preserved food and the lecithinase activity of isolated strains].

This is a study of 414 different foodstuff types for Cl. perfringens presence determination. Of the meat products are with the highest contamination - 32 per cent, next ranking seasoning spices ond condiments, and auxiliary materials - 10.9 per cent. In milk, dairy and confectionary products it is isolated in 3.8 per cent of samples, while in tinned foodstuffs - in 1.8 per cent. Lecithinase production of the isolated strains in Kitt-Tarozi bouillon, Following 18-20 hour-long cultivation at 37 degrees C is studied, and dosed within limits ranging from 5 to 100 DMO. In meat food - pork in its own sauce, the development and lecithinase activity of some strains with different doses lecithinase in the enriching bouillon is studied after cultivation of the same at room temperature for 18-20 hours. Under the conditions outlined it is established that the growth of the strains is analogical to that of the bouillon, whilst their lecithinase activity shows a several-fold increase, and is dependent on the individual characteristics of the strains, and on their adaptive properties. The produced quantity of lecithinase does not affect the toxicity of strains upon oral infection of the animals experimented.

Animals

Investigations of source and route of Yersinia enterocolitica infection.

Isolation of Yersinia enterocolitica was performed on healthy humans, animals and food, and growth of Y. enterocolitica on sliced ham at various temperature ranges was also examined. It was noticed that the most prevalent 0 group (03) in human infection was isolated from swine, pork and chopping boards. The storage of food in the refrigerator (5 degrees C) possibly increases the risk of infection since Y. enterocolitica is multipliable at low temperatures more readily than many other pathogenic bacteria. Pork and meat products contaminated with Y. enterocolitica, either primarily or secondarily, are probably the most important source of human infection.

Adult

Sylvatic trichinosis in British Columbia: potential threat to human health from an independent cycle.

The results of a 3-year study of trichinosis in British Columbia wild-life, based on the testing of more than 9,000 tissue specimens from a large variety of animal species, indicated that trichinosis is widespread among wild mammals in the southern and central parts of British Columbia. This continuing survey has established that the disease is carried by at least 15 species of terrestrial mammals including 3 species of rodents. The finding of Trichinella spiralis in ground squirrels and nonsynanthropic mice may be the first reported in North America. Although trichinosis appears to be eradicated in domestic pigs in British Columbia, a sylvatic cycle of the disease continues to exist independently and poses a potential threat to human health. It is possible for human beings to contract trichinosis by consuming inadequately cooked meat from certain wildlife species, especially bears, as well as meat products (such as pork or beef sausage) to which game meat has been added; several local outbreaks were caused by this source. Another hazard of unknown potential is the spread of trichinosis from the wild animal reservoir--from rodents in particular--to domestic pigs and thus to man.

Animals

[Residues of organochlorine insecticides and PCBs in some Norweigian foods (author's transl)].

Oats, barley, wheat and rye, meat from sheep, pork, beef and chicken/broiler, boullion, margarine and cod roe were analysed by gas liquid chromatography for chlorimated insecticides and PCBs. Residues of PCBs, lindane, DDT and metabolites were found in only a few of the samples. Traces of dieldrin were observed in grain. Lidan was found in grain with the highest single value of 0.013 ppm and with the mean value of 0.008 ppm. DDT residues were found in 37 percent of all samples and the highest concentration were in chicken/broiler with mean value of 0.02 ppm. The values are calculated on wet weight basis.

Animals

[Determination of connective tissue content of meat products using the hydroxyproline methods (author's transl)].

The amino acids of collagenous and elastic connective tissue of beef and pork and of commercial meat products were determined by gaschromatography. A correlation was established between hydroxyproline and glycine and also between hydroxyproline and proline. Thus the content of elastic connective tissue in meat products is quite constant and compared with the amount of collagenous material insignificant.--Determination of amount of connective tissue in meat products from the amount of hydroxyproline this is suitable for the needs of food control.

Amino Acids

Nutrient levels of some foods of Eskimos from Artic Bay, N. W.T., Canada.

Native food samples of caribou, seal, and arctic char were collected from resident Eskimos in Arctic Bay, N.W. T., and analyzed for nutrients. Caribou and seal meats were similar in protein and fat content in comparison with beef. Arctic char was above the average in protein and fat content for fish. Ascorbic acid was present in significant amounts only in baby seal liver. Thiamin, riboflavin, vitamin B12, folacin, and pantothenic acid content ranged from average to high for seal and carbiou meat, respectively, in comparison to published values for pork and beef. The vitamin B6 content of caribou meat was comparable to average values reported for pork and beef but was lower in seal meat. Baby seal liver contained similar levels of thiamin, riboflavin, vitamin B12, folacin, and pantothenic acid and less vitamin B6 than those reported for pork, beef, calf, and lamb liver. Arctic char was not appreciably different in these vitamins from other fish. The iron content of the Eskimo foods was higher than the meat group of the mixed Canadian diet. The macro mineral nutrients in these foods were comparable to those in beef and fish. The amino acid composition of caribou and seal meat was similar to beef, except that seal meat had a lower content of sulfur-containing amino acids and a much higher content of histidine. The sample of seal and arctic char were relatively high in concentrations of long-chain unsaturated fatty acids.

Amino Acids

[Salmonella in minced meat from ten meat inspection services in the Netherlands (author's transl)].

Throughout a period of thirteen months, samples of minced meat from ten different meat inspection services in the Netherlands were examined weekly for the presence of Salmonella. One hundred samples of 150 g. of each meat inspection service were studied by six different methods. Marked differences in contamination of minced meat with Salmonella were observed between the various localities. These were differences in the proportion of contaminated samples from each meat inspection service (lowest 10%, highest 39%) on the one hand and differences in the number of contaminated sub-samples of each sample of minced meat (smallest 18, largest 139) on the other. Moreover, the season obviously was a factor in contamination of minced meat with Salmonella, the contamination rate increasing with the outside temperature. As a rule, contamination by Salmonella was less common in minced beef than it was in minced beef and pork mixed. Contamination of minced meat with Salmonella usually was less common in minced meat from butchers' shops in which slaughtering was done by the butcher himself than it was in minced meat from slaughterhouses. The organisms most frequently isolated in the present study were S. typhi murium phage type II 505, S. panama and S. brandenburg. Of the methods of isolation used, those with pre enrichment and direct enrichment in TBB at 43 degrees C were found to produce the most satisfactory results.

Abattoirs

The detection of chicken meat in meat products by means of the anserine/carnosine ratio.

A distinctive difference was found between the ratio of the anserine and carnosine contents (a/c ratio) in beef or pork and of that in chicken/meat. The a/c ratio for beef varies between 0.06-0.2 and for pork between 0.02-0.1 but for chicken meat can reach values as high as 2.2-5.5. The high a/c ratio for chicken meat proved to be sufficient to detect this ingredient at a 5% level in both cooked pork products and 1:1 beef-pork mixtures, this being independent of the heat treatment. The a/c ratio should be considered to be a suitable parameter for the presence of chicken meat in meat products.

Animals

Cooking frozen and thawed roasts: beef, pork, and lamb cuts.

Cooking time, yield, and palatability of paired beef, pork, and lamb roasts cooked from the frozen and thawed states were compared. Cooking time for all roasts averaged from 3 to 22 min. per pound longer for meat cooked from the frozen state. The longer cooking time from the frozen state. The longer cooking time from the frozen state was greater for roasts with a large amount of bone and for cuts cooked by braising than for less bony roasts and cuts cooked by roasting. Except for thawed beef rump roasts, which had a higher yield of cooked lean meat, yield of cooked lean meat from the various cuts of beef, pork, and lamb was not affected by the state at the start of cooking. Collectively, all pork roasts had a higher yield of cooked lean meat when cooked from the frozen state. Juiciness and natural flavor of the roasts were not affected by the state at the start of cooking. Lamb leg and rib roasts were more tender when cooked from the thawed state.

Animals

Outbreak of toxoplasmosis in a family and documentation of acquired retinochoroiditis.

An outbreak of toxoplasmosis in one household is described. It demonstrates the potential for a common source infection with Toxoplasma gondii to cause multiple cases. Six of seven members of a household investigated for toxoplasmosis demonstrated high antibody titers consistent with recent infection; five of these members (83%) were symptomatic. The most common manifestations were fever and lymphadenopathy, which developed from seven to 18 days (mean 11 days) after a common source ingestion of infected meat. Since inadequately cooked lamb, pork and beef are probably the most common sources of infection in the United States, outbreaks of multiple cases may occur more frequently than is generally appreciated. As more outbreaks of febrile illnesses are examined, especially in families and closed communities, it is likely that more common source epidemics of toxoplasmosis will be recognized. Retinochoroiditis is an unusual manifestation of adult acquired toxoplasmosis. In the index case in this epidemic the patient manifested vision threatening retinochoroiditis 129 days after infection with toxoplasmosis. He represents the ninth well-documented case of toxoplasma retinochoroiditis associated with adult acquired disease.

Adolescent